@article {pmid40397277,
year = {2026},
author = {Lodewijk, GA and Kozuki, S and Guiltinan, C and Topacio, BR and Shariati, SA},
title = {Application of CRISPR-Based Epigenome Editing Tools for Engineering Programmable Embryo Models.},
journal = {Methods in molecular biology (Clifton, N.J.)},
volume = {3048},
number = {},
pages = {211-238},
pmid = {40397277},
issn = {1940-6029},
support = {R35 GM147395/GM/NIGMS NIH HHS/United States ; },
mesh = {*Epigenome Editing/methods ; Animals ; Mice ; *CRISPR-Cas Systems ; *Embryonic Development/genetics ; Humans ; *Embryo, Mammalian/cytology ; Embryonic Stem Cells/cytology/metabolism ; },
abstract = {Stem cell-based embryo models (SEMs) have the potential to transform our understanding of early human embryogenesis. A critical step in engineering SEMs is the generation of the major cell types that compose preimplantation embryos including two primary extraembryonic lineages: (i) trophoblast cells, which are crucial for implantation and the establishment of maternal-fetal exchange, and (ii) hypoblast cells, which contribute to yolk sac formation. In addition, both cell types provide key signaling cues necessary for embryonic development. CRISPR-based epigenome editors are programmable devices that allow for efficient and precise activation (CRISPRa) or repression (CRISPRi) of cell fate-determining factors by modulating endogenous regulatory elements. Here, we present a step-by-step method to implement CRISPRa for controlling cell fate in embryonic stem cells based on our work in generation of CRISPR-programmed mouse embryo models.},
}

*Epigenome Editing/methods
Animals
Mice
*CRISPR-Cas Systems
*Embryonic Development/genetics
Humans
*Embryo, Mammalian/cytology
Embryonic Stem Cells/cytology/metabolism

@article {pmid42218379,
year = {2026},
author = {Shi, Y and Yin, J and Ning, S and Yuan, J and Yang, D and Chuai, G},
title = {DeepCas12a: a hybrid deep learning framework for accurate AsCas12a efficiency prediction from sequence and epigenetic information.},
journal = {BMC genomics},
volume = {27},
number = {1},
pages = {},
pmid = {42218379},
issn = {1471-2164},
support = {2025080107//Tongji University "Medicine + X" Cross Research Program/ ; 62002265//National Natural Science Foundation of China/ ; },
mesh = {*Deep Learning ; *Epigenesis, Genetic ; *CRISPR-Cas Systems ; Convolutional Neural Networks ; *Gene Editing/methods ; *CRISPR-Associated Proteins/genetics/metabolism ; Bacterial Proteins ; Endodeoxyribonucleases ; },
abstract = {CRISPR-Cas12a (Cpf1) offers distinct advantages for genome editing due to its flexible, T-rich PAM recognition. However, variable cleavage efficiency-modulated by sequence context and epigenetic features-remains a challenge, with existing tools facing challenges in modeling the high-order interactions between multimodal features. Here, we present DeepCas12a, a hybrid deep learning framework integrating Convolutional Neural Networks (CNNs) and a Vision Transformer (ViT) encoder to capture both local sequence motifs and long-range dependencies. The model fuses DNA sequence data with epigenetic profiles (DNA methylation and chromatin accessibility) in an end-to-end architecture. Benchmarked on an independent test set, DeepCas12a outperformed state-of-the-art predictors, achieving an Average Precision of 0.783, an AUC of 0.868, and a Spearman correlation of 0.630. Furthermore, interpretability analysis via saliency maps confirms the model captures biologically relevant features, including PAM specificity and seed region sensitivity, facilitating rational guide RNA design.},
}

*Deep Learning
*Epigenesis, Genetic
*CRISPR-Cas Systems
Convolutional Neural Networks
*Gene Editing/methods
*CRISPR-Associated Proteins/genetics/metabolism
Bacterial Proteins
Endodeoxyribonucleases

@article {pmid42350674,
year = {2026},
author = {Navalayeu, T and Beer, N and Bebjaková, M and Kalis, RW and Hohmann, U and Stejskal, K and Krššáková, G and Fasching, N and Herzog, VA and Popitsch, N and Roitinger, E and Plaschka, C and Zuber, J and Ameres, SL},
title = {Cellular assembly and functional resilience of the mammalian RNA exosome.},
journal = {The EMBO journal},
volume = {45},
number = {15},
pages = {5423-5456},
pmid = {42350674},
issn = {1460-2075},
support = {LS23-053//Vienna Science and Technology Fund (WWTF)/ ; CoG-866166//EC | European Research Council (ERC)/ ; 10.55776/F80//Austrian Science Fund (FWF)/ ; 10.55776/DOC177//Austrian Science Fund (FWF)/ ; PhD Fellowship//Boehringer Ingelheim Fonds (BIF)/ ; },
mesh = {Animals ; Mice ; *Exosome Multienzyme Ribonuclease Complex/metabolism/genetics ; *Exosomes/metabolism ; RNA-Binding Proteins/metabolism/genetics ; Proteasome Endopeptidase Complex/metabolism ; *Mouse Embryonic Stem Cells/metabolism ; CRISPR-Cas Systems ; RNA/metabolism ; },
abstract = {Most eukaryotic proteins assemble into multisubunit complexes that coordinate essential cellular functions, yet the principles governing their assembly and proteostatic control remain largely undefined. Here, we systematically dissect the cellular assembly and functional organization of the RNA exosome, an essential ribonucleolytic complex, using an inducible dual-guide CRISPR/Cas9 system in mouse embryonic stem cells. We reveal a sequential assembly pathway where Exosc2, Exosc4, and Exosc7 initiate complex formation, facilitating the incorporation of barrel and cap subunits in a defined hierarchy. Unlike other structural subunits, the terminally incorporated cap subunit Exosc1 is dispensable for cell viability, revealing a modular, functionally resilient architecture. We demonstrate that orphan subunits are selectively degraded via the ubiquitin-proteasome system, enforcing stringent quality control over RNA exosome biogenesis. These findings define an assembly logic of of the mammalian exosome and uncover previously unrecognized plasticity in the composition and function of this essential ribonucleolytic complex.},
}

Animals
Mice
*Exosome Multienzyme Ribonuclease Complex/metabolism/genetics
*Exosomes/metabolism
RNA-Binding Proteins/metabolism/genetics
Proteasome Endopeptidase Complex/metabolism
*Mouse Embryonic Stem Cells/metabolism
CRISPR-Cas Systems
RNA/metabolism

@article {pmid42462663,
year = {2026},
author = {Chen, H and Jin, Z and Duan, C and Song, Z and Cheng, Y and Chen, M and Zhang, C and Lan, Y and Shen, W and Fu, Y and Liu, R and Zheng, X},
title = {Effects of the ecpA gene on the biological characteristics and pathogenicity of avian pathogenic Escherichia coli strain FJLY68.},
journal = {Veterinary microbiology},
volume = {320},
number = {},
pages = {111145},
doi = {10.1016/j.vetmic.2026.111145},
pmid = {42462663},
issn = {1873-2542},
mesh = {Animals ; *Escherichia coli/pathogenicity/genetics ; *Escherichia coli Infections/microbiology/veterinary ; *Poultry Diseases/microbiology ; Chickens/microbiology ; Biofilms/growth & development ; Virulence/genetics ; *Escherichia coli Proteins/genetics/metabolism ; Bacterial Adhesion/genetics ; Fimbriae, Bacterial/genetics ; Chick Embryo ; *Fimbriae Proteins/genetics ; Cell Line ; CRISPR-Cas Systems ; Genetic Complementation Test ; Gene Deletion ; },
abstract = {Avian pathogenic Escherichia coli (APEC) is a major cause of colibacillosis in poultry, yet the role of the ecpA gene, which encodes the major structural subunit of the Escherichia coli common pilus (ECP), remains incompletely defined in APEC pathogenesis. To investigate the role of ecpA in the biological characteristics and pathogenicity of Avian Pathogenic Escherichia coli (APEC) strain FJLY68, an ecpA deletion mutant (ΔecpA) and its corresponding complemented strain (CΔecpA) were constructed using the CRISPR/Cas9 system and verified by PCR and Sanger sequencing. Phenotypic analyses revealed that the ΔecpA mutation significantly impaired bacterial motility, biofilm formation, adherence to chicken embryonic fibroblast (DF-1) cells, and fimbriae assembly. Transcriptomic analysis identified 1720 differentially expressed genes in the ΔecpA mutant, significantly enriched in pathways associated with flagellar assembly, chemotaxis, and metabolism, consistent with the observed phenotypic changes. Although in vitro growth was unaffected, the ΔecpA mutant exhibited markedly attenuated virulence in a chick infection model, as indicated by an increased LD50, attenuated clinical signs and pathological lesions, and reduced bacterial colonisation in tissues. Full genetic complementation restored all observed defects to wild-type levels. This study identifies ecpA as a critical determinant of APEC pathogenesis, directly linking its function to bacterial motility, biofilm formation, adhesion, and in vivo virulence, and provides a theoretical basis for developing novel control strategies targeting this virulence factor.},
}

Animals
*Escherichia coli/pathogenicity/genetics
*Escherichia coli Infections/microbiology/veterinary
*Poultry Diseases/microbiology
Chickens/microbiology
Biofilms/growth & development
Virulence/genetics
*Escherichia coli Proteins/genetics/metabolism
Bacterial Adhesion/genetics
Fimbriae, Bacterial/genetics
Chick Embryo
*Fimbriae Proteins/genetics
Cell Line
CRISPR-Cas Systems
Genetic Complementation Test
Gene Deletion

@article {pmid42470400,
year = {2026},
author = {Ang, YS and Yung, LL},
title = {Sequence Engineering of Guide DNA for Precise RNA Targeting by Cas12a.},
journal = {ACS nano},
volume = {20},
number = {30},
pages = {21115-21127},
doi = {10.1021/acsnano.6c02663},
pmid = {42470400},
issn = {1936-086X},
support = {NA//Singapore Ministry of Education Academic Research Fund Tier 1/ ; NA//Singapore Ministry of Health?s National Medical Research Council, Programme for Research in Epidemic Preparedness and Response (PREPARE)/ ; },
mesh = {*CRISPR-Associated Proteins/metabolism/chemistry/genetics ; *DNA/chemistry/genetics/metabolism ; *Endodeoxyribonucleases/metabolism/chemistry/genetics ; *Bacterial Proteins/metabolism/genetics/chemistry ; *RNA/metabolism/chemistry/genetics ; *RNA, Guide, CRISPR-Cas Systems/genetics ; Base Sequence ; },
abstract = {Cas12a is highly accommodative toward noncanonical activation pathways to the extent of flipping its identity to be a DNA-guided RNA-targeting effector. A sequence engineering approach was used to systematically identify desirable guide DNA (gDNA) sequence motifs to achieve comparable RNA targeting efficiency as the canonical RNA-guided Cas12a with good selectivity down to single-nucleotide mismatch. Importantly, we introduced a split gDNA design concept with greater energetic differences arising from subtle nucleotide changes to probe the key spacer features for effective Cas12a-gDNA activation. Similar to the canonical RNA-guided activation pathway, Cas12a was found to engage actively in the "seed-like" scaffold-proximal region while the scaffold-distal region was largely hybridization-driven. We further evolved the split gDNA design to enhance the sequence selectivity by up to 21-fold compared to a single gDNA design and achieve single-nucleotide discrimination among representative let-7 family members. This study has established a gDNA sequence design framework to reprogram Cas12a as a precise RNA targeting platform.},
}

*CRISPR-Associated Proteins/metabolism/chemistry/genetics
*DNA/chemistry/genetics/metabolism
*Endodeoxyribonucleases/metabolism/chemistry/genetics
*Bacterial Proteins/metabolism/genetics/chemistry
*RNA/metabolism/chemistry/genetics
*RNA, Guide, CRISPR-Cas Systems/genetics
Base Sequence

@article {pmid42480873,
year = {2026},
author = {McClain, IM and Yigit, NS and Royzen, M},
title = {Non-chromatographic purification of guide RNA for gene-editing experiments.},
journal = {Bioorganic & medicinal chemistry letters},
volume = {140},
number = {},
pages = {130740},
doi = {10.1016/j.bmcl.2026.130740},
pmid = {42480873},
issn = {1464-3405},
mesh = {Humans ; *RNA, Guide, CRISPR-Cas Systems/isolation & purification/genetics/chemistry ; *Gene Editing/methods ; CRISPR-Cas Systems ; Chromatography, High Pressure Liquid ; Green Fluorescent Proteins/genetics ; },
abstract = {CRISPR-Cas12a gene editing technology is gaining momentum as a powerful tool for many biochemical and medicinal applications. The technology requires guide RNA, which is typically made using solid phase synthesis and purified by HPLC. The latter is often the most complex and time-consuming element of the synthetic process. This communication describes a non-chromatographic method for purification of synthetic RNAs. The method consists of five steps and yields target RNA in over 80% purity, which adheres to the FDA's standard for gene editing applications. The non-chromatographic RNA purification approach was applied to synthesize guide RNA targeting the GFP gene. Its purity was analyzed by analytical HPLC. Its functional fidelity was tested in CRISPR-Cas12a experiments in solution and live mammalian cells.},
}

Humans
*RNA, Guide, CRISPR-Cas Systems/isolation & purification/genetics/chemistry
*Gene Editing/methods
CRISPR-Cas Systems
Chromatography, High Pressure Liquid
Green Fluorescent Proteins/genetics

@article {pmid42518166,
year = {2026},
author = {Kang, J and Kim, HH and Yoon, HM and Choi, Y and Heo, J and Woo, Y and Yu, S and Lee, KH and Lee, Y},
title = {Phenotypic and Whole-Genome Characterization of Enterococcus Isolates from Korean Doenjang and Meju: E. durans Edu-1 as a Food-Grade Probiotic Candidate with Epithelial Wound-Healing Activity.},
journal = {Probiotics and antimicrobial proteins},
volume = {},
number = {},
pages = {},
pmid = {42518166},
issn = {1867-1314},
support = {2026-RISE-09-A32//the Regional Innovation System & Education (RISE) program through the Gyeonggi Province RISE initiative, funded by the Ministry of Education (MOE) and the Gyeonggi Province, the Republic of Korea/ ; },
abstract = {Enterococcus species are widely distributed in traditional fermented foods and have shown probiotic potential. However, their food-grade application has been limited because the European Food Safety Authority excluded the genus from the qualified presumption of safety (QPS) list. Therefore, strain-specific safety assessment is essential. In this study, we characterized Enterococcus isolates from Korean Doenjang and Meju using an integrated phenotypic and whole-genome sequencing approach. Sixteen strains were isolated and identified/determined by 16S rRNA gene sequencing. We evaluated phenotypic safety and probiotic traits, and five candidates were further tested in HT-29 wound-healing scratch assays and a 115-gene host expression array. The two best wound-healing strains were then analyzed by whole-genome sequencing using a multi-database approach (VirulenceFinder, VFDB, ResFinder, CARD/RGI, PlasmidFinder, CRISPRCasFinder, and antiSMASH). The 16 isolates belonged to the five species, among which E. durans Edu-1 showed the highest wound closure ability (18.75 ± 1.24% at 72 h), followed by E. raffinosus Era-1 (16.71 ± 0.49%). Whole-genome sequence analysis showed that Edu-1 had no acquired transmissible antibiotic resistance and virulence factors genes and in addition, contains a plasmid-borne Bacteriocin_II family biosynthetic gene cluster, which can explain/support its broad-spectrum antimicrobial activity. In contrast, Era-1 carried tet(M) gene on the chromosome and ermB gene on an insertion-sequence-rich mobile element, although it also has a functional Type I-B CRISPR-Cas system and three glutamate decarboxylase (gadB) genes for γ-aminobutyric acid biosynthesis. These findings identify/suggest E. durans Edu-1 as a food-grade probiotic candidate with epithelial wound-healing activity, while E. raffinosus Era-1 represents a strain of biological interest for mechanistic wound-healing research rather than food application.},
}

@article {pmid42519596,
year = {2026},
author = {Hess, WR and Marchfelder, A and Randau, L},
title = {Editorial: creative CRISPR-Cas: RNA-guided functions in defence and beyond.},
journal = {microLife},
volume = {7},
number = {},
pages = {uqag026},
pmid = {42519596},
issn = {2633-6693},
}

@article {pmid42521019,
year = {2026},
author = {Dalabehera, M and Chaudhari, S and Kumar, J and Poonia, N and Subudhi, RN and Choonara, YE and Figueiras, A and Shukla, N and Sharma, N and Mascarenhas-Melo, F},
title = {Mechanistic advances in nanomedicine, nucleic acid therapies, and AI-driven research on cervical cancer.},
journal = {Journal of pharmaceutical sciences},
volume = {},
number = {},
pages = {104442},
doi = {10.1016/j.xphs.2026.104442},
pmid = {42521019},
issn = {1520-6017},
abstract = {Cervical cancer pharmacotherapy is significantly limited by physiological and cellular barriers that restrict drug access to therapeutic targets, resulting in suboptimal biodistribution, systemic toxicity, and the emergence of drug resistance. This review provides a mechanistic and biopharmaceutics-centered analysis of how advanced drug delivery systems are being engineered to overcome these limitations. We critically examine the role of nanocarriers, including lipid-based vesicles, polymeric nanoparticles, and inorganic hybrid systems, in modulating absorption, distribution, and tumor-targeting efficiency, with emphasis on their physicochemical properties and interaction with biological barriers such as the tumor microenvironment and cellular uptake pathways. In parallel, we analyze nucleic acid-based therapeutics (CRISPR/Cas systems, miRNA, and antisense oligonucleotides) from a pharmaceutical sciences perspective, focusing on delivery constraints, stability, intracellular trafficking, and their ability to modulate pharmacological response and drug resistance mechanisms. The review also discusses the integration of immunomodulatory strategies within nanodelivery platforms as a means to alter disease-related biological barriers and improve therapeutic index. Finally, we explore the emerging role of AI-assisted models in optimizing formulation design, predicting pharmacokinetic behavior, and supporting precision dosing strategies in drug development workflows. By integrating drug delivery engineering, molecular biopharmaceutics, and computational optimization, this work outlines a translational framework for overcoming key barriers in pharmaceutical intervention design for oncology applications.},
}

@article {pmid42521788,
year = {2026},
author = {Liu, H and Yuan, Z and Han, J and Zhao, Y},
title = {A dual-readout RAA-CRISPR/Cas13a diagnostic platform for rapid and sensitive detection of Eggerthella lenta.},
journal = {Mikrochimica acta},
volume = {193},
number = {8},
pages = {},
pmid = {42521788},
issn = {1436-5073},
support = {2019LJ001//Academic Promotion Program of Shandong First Medical University/ ; 2021ZDSYS27//the Key Research and Development Project of Shandong Province/ ; },
mesh = {*CRISPR-Cas Systems ; *Actinobacteria/isolation & purification/genetics ; Limit of Detection ; Humans ; *Nucleic Acid Amplification Techniques/methods ; Rapid Diagnostic Tests ; Recombinases/metabolism ; },
abstract = {Eggerthella lenta (E. lenta) is an opportunistic anaerobic pathogen associated with severe systemic infections, yet rapid and accurate diagnostic tools remain limited. To address this challenge, we developed a highly sensitive and specific dual-readout diagnostic platform integrating recombinase-aided amplification (RAA) with the CRISPR/Cas13a system, targeting the highly conserved rsmG gene of E. lenta. The assay offers two detection modalities: a real-time fluorescence readout and a visually interpretable lateral flow strip. Analytical evaluation demonstrated that the fluorescence-based assay achieved a limit of detection (LOD) of 4.4 copies per reaction (95% CI: 3.7-5.6 copies/reaction), while the instrument-free lateral flow assay yielded an LOD of 10[4] copies per reaction. The platform exhibited exceptional specificity, showing no cross-reactivity with 10 common non-target bacterial species. Clinical validation was performed using 24 synovial fluid samples, all confirmed positive for E. lenta by Sanger sequencing. The fluorescence assay successfully detected all 24 samples, achieving a detection rate of 100% (24/24). In parallel, the lateral flow assay detected 21 of the 24 positive samples, yielding a detection rate of 87.5% (21/24). The three samples undetected by the lateral flow strip were verified as true positives by sequencing, indicating that the discrepancy was due to the lower analytical sensitivity of the strip format rather than a lack of specificity. In conclusion, this dual-mode RAA-CRISPR/Cas13a platform serves as a robust and practical tool for rapid clinical diagnosis and point-of-care (POC) triaging of E. lenta infections. The fluorescence format is optimal for high-sensitivity laboratory testing, whereas the lateral flow variant provides a deployable alternative for rapid, point-of-care screening in resource-limited environments.},
}

*CRISPR-Cas Systems
*Actinobacteria/isolation & purification/genetics
Limit of Detection
Humans
*Nucleic Acid Amplification Techniques/methods
Rapid Diagnostic Tests
Recombinases/metabolism

@article {pmid42521823,
year = {2026},
author = {Tuo, W and Wang, X and Wu, T and Zhang, S and Guo, P and Lin, X and Lin, Q and Zhai, J and Wu, S},
title = {Establishment of an efficient Agrobacterium-mediated transformation system and CRISPR/Cas9-mediated genome editing of the OvPDS1 gene in Oxalis vulcanicola 'Sunset Velvet'.},
journal = {Planta},
volume = {264},
number = {3},
pages = {},
pmid = {42521823},
issn = {1432-2048},
support = {32471958//National Natural Science Foundation of China/ ; KFB24022A//Study on the Molecular Mechanisms of Leaf Coloration in Three Sulfur Oxalis Varieties, Innovation Project of Fujian Agriculture and Forestry University/ ; },
mesh = {*CRISPR-Cas Systems/genetics ; Plants, Genetically Modified/genetics ; *Transformation, Genetic ; *Gene Editing/methods ; *Agrobacterium/genetics ; *Plant Proteins/genetics/metabolism ; Acetophenones ; },
abstract = {This study overcomes a key technical barrier by establishing transformation and enabling first CRISPR/Cas9 editing in Oxalis, providing a platform for functional genomics and breeding. The lack of an efficient genetic transformation system has considerably hindered functional genomics studies in Oxalis vulcanicola 'Sunset Velvet'. Here, we established a stable and efficient Agrobacterium-mediated transformation system using stem segments as explants. Key parameters, including pre-culture duration, infection time, Agrobacterium cell density, acetosyringone (AS) concentration, and co-cultivation period, were systematically optimized. Under optimal conditions, the highest transient β-glucuronidase (GUS) expression rate reached approximately 9.0%, and eight stable transgenic lines were successfully obtained. CRISPR/Cas9-mediated genome editing was achieved in Oxalis for the first time. Targeted mutagenesis of OvPDS1, a gene involved in carotenoid biosynthesis, resulted in an albino phenotype, and Sanger sequencing confirmed a base substitution at the target site. Although the editing efficiency was relatively low (0.5%), this result demonstrates the feasibility of genome editing in Oxalis. This study overcomes a major technical bottleneck and provides a robust platform for functional gene analysis, trait improvement, and molecular breeding in O. vulcanicola 'Sunset Velvet' and other non-model ornamental plants.},
}

*CRISPR-Cas Systems/genetics
Plants, Genetically Modified/genetics
*Transformation, Genetic
*Gene Editing/methods
*Agrobacterium/genetics
*Plant Proteins/genetics/metabolism
Acetophenones

@article {pmid42522380,
year = {2026},
author = {Xue, F and Xin, Z and Wang, G and Xing, J and Han, H and Zhao, X and Song, N},
title = {Recent advances of CRISPR-based gene editing technologies and delivery strategies.},
journal = {Artificial cells, nanomedicine, and biotechnology},
volume = {54},
number = {1},
pages = {415-431},
doi = {10.1080/21691401.2026.2702869},
pmid = {42522380},
issn = {2169-141X},
mesh = {Humans ; *Gene Editing/methods ; Animals ; *CRISPR-Cas Systems/genetics ; *Gene Transfer Techniques ; },
abstract = {CRISPR technology is a powerful tool for gene editing, in which the efficient delivery of living target cells allows it to show great clinical potential. At present, the commonly used in vivo delivery strategies mainly include biological methods (AAV, VLP, SEND) and chemical methods (LNP), which subtly deliver gene editors to living target cells safely and efficiently from different ways. However, existing delivery systems have different extents of limitations in terms of editing efficiency, immunogenicity, half-life, etc., so developing optimized delivery systems is the key to fully realizing the potential of CRISPR-Cas system for intracellular gene editing. In order to fully understand the advantages of different delivery strategies to maximize the ability to help CRISPR systems choose delivery methods, we conducted a systematic review. In this paper, we introduce the types, principles and characteristics of gene editing systems in order to understand their requirements for delivery tools. We focus on describing the type, principle, load, immunogenicity, specificity, toxicity, etc. of the delivery system, so as to fully analyse its advantages and disadvantages for the selection of different editing environments. This review aims to provide new insights to facilitate appropriate delivery systems or improve the efficacy of existing systems.},
}

Humans
*Gene Editing/methods
Animals
*CRISPR-Cas Systems/genetics
*Gene Transfer Techniques

@article {pmid42522597,
year = {2026},
author = {Lu, W and Yang, Q and Zhao, P and Cao, Y and Jing, Z and Zhang, N and Li, J and Li, X and Wang, X and Zhang, X and Cao, L and Gong, P},
title = {A One-Pot RPA-CRISPR/Cas12a Assay for Rapid Genus-Level Detection of Babesia spp. in Ticks and Livestock Blood Samples.},
journal = {Transboundary and emerging diseases},
volume = {2026},
number = {1},
pages = {e9289663},
pmid = {42522597},
issn = {1865-1682},
support = {2024YFD1800100//National Key Research and Development Program of China/ ; CARS-39//China Wool-sheep & Cashmere-goat Research System/ ; },
mesh = {Animals ; *Babesia/isolation & purification/genetics ; *Babesiosis/diagnosis/parasitology/blood ; Cattle ; *Cattle Diseases/diagnosis/parasitology/blood ; *Sheep Diseases/diagnosis/parasitology/blood ; CRISPR-Cas Systems ; *Ticks/parasitology ; Sheep ; *Nucleic Acid Amplification Techniques/veterinary/methods ; Rapid Diagnostic Tests ; Sensitivity and Specificity ; Livestock ; Clustered Regularly Interspaced Short Palindromic Repeats ; },
abstract = {Babesiosis, a globally significant tick-borne disease, poses substantial threats to livestock production and public health. Reported cases of human babesiosis in the United States increased from 1742 in 2014 to 3586 in 2023. In livestock, cattle babesiosis causes mortality, reduced meat and milk production, reproductive losses, and substantial control costs, with annual economic losses estimated at hundreds of millions of US dollars in several endemic countries. Rapid and sensitive detection methods are essential for early warning, surveillance, and control of this disease. In this study, we developed a closed-tube, one-pot assay for genus-level detection of Babesia spp. associated with cattle and sheep, based on recombinase polymerase amplification (RPA) coupled with clustered regularly interspaced short palindromic repeats (CRISPR)/Cas12a. This format effectively minimizes cross-contamination risks associated with repeated tube opening in conventional assays. A three-channel signal readout system, including blue-light fluorescence visualization, ultraviolet (UV) fluorescence visualization, and lateral flow strip (LFS) readout, was integrated to enable flexible endpoint detection under different laboratory and field conditions. The assay targets a conserved region of the Babesia 18S rRNA gene and enables genus-level detection of Babesia spp. within 40 min at 37°C. The established RPA-CRISPR/Cas12a platform exhibited high analytical sensitivity, with a limit of detection of 5 copies/μL for recombinant plasmid templates, high analytical specificity against the tested nontarget pathogens, and low equipment dependency. The detection limit of the LFS format reached 50 copies/μL. Field validation using 71 pooled tick samples and 53 clinical blood samples collected from cattle and sheep yielded positive rates of 15.49% and 9.43%, respectively, with 100% concordance between this assay and conventional polymerase chain reaction (PCR) for both specimen types. In conclusion, this one-pot RPA-CRISPR/Cas12a detection platform provides a rapid, sensitive, and field-applicable molecular screening tool for genus-level detection of Babesia spp. This assay may support early warning and preliminary field monitoring of babesiosis, particularly in resource-limited settings. However, species-level confirmation should be performed by sequencing or other species-specific methods when epidemiological tracing or precise species identification is required.},
}

Animals
*Babesia/isolation & purification/genetics
*Babesiosis/diagnosis/parasitology/blood
Cattle
*Cattle Diseases/diagnosis/parasitology/blood
*Sheep Diseases/diagnosis/parasitology/blood
CRISPR-Cas Systems
*Ticks/parasitology
Sheep
*Nucleic Acid Amplification Techniques/veterinary/methods
Rapid Diagnostic Tests
Sensitivity and Specificity
Livestock
Clustered Regularly Interspaced Short Palindromic Repeats

@article {pmid42524122,
year = {2026},
author = {Rananaware, SR and Narisetty, KV and Shah, RA and Jain, PK},
title = {CRISPR-based ex vivo gene editing of donor organs.},
journal = {Nature reviews bioengineering},
volume = {},
number = {},
pages = {},
pmid = {42524122},
issn = {2731-6092},
support = {R21 AI156321/AI/NIAID NIH HHS/United States ; R21 AI168795/AI/NIAID NIH HHS/United States ; R35 GM147788/GM/NIGMS NIH HHS/United States ; R61 AI181016/AI/NIAID NIH HHS/United States ; },
abstract = {Donor organs are frequently discarded because of concerns about quality or pathogen risk, challenges that could be mitigated through ex vivo gene editing or silencing during machine perfusion. Here, we discuss the development of CRISPR-based approaches for ex vivo gene silencing in human donor organs, from proof of concept in kidney biopsies to the challenges of organ-scale translation.},
}

@article {pmid42524609,
year = {2026},
author = {Martin, L and Bohinc, J and Recchia, A and Gritti, S and Santilli, G and Zeyland, J and Vidaković, M and Grdović, N and Benabdellah, K and Ortiz-Bueno, M and Butuner, BD and Pisaniello, L and Stilhano, R and Benati, D and Hapil, FZ and Khawaja, S and Nair, RR and Giacomelli, C and Atilla, E and Zinghirino, F and Ferrari, T and Corradi, F and Laufer, TJ and Khnykin, D and Aseguinolaza, GG and Skrbinek, M and Mlakar, T and Lapanja, T and Lainšček, D},
title = {In vivo delivery strategies for therapeutic CRISPR genome editing.},
journal = {International journal of biological sciences},
volume = {22},
number = {12},
pages = {6539-6581},
pmid = {42524609},
issn = {1449-2288},
mesh = {Humans ; *Gene Editing/methods ; Animals ; *CRISPR-Cas Systems/genetics ; Epigenome Editing ; Genetic Therapy/methods ; *Gene Transfer Techniques ; },
abstract = {CRISPR-based genome and epigenome editing technologies have rapidly evolved from programmable nucleases into a diverse therapeutic toolbox encompassing conventional CRISPR systems, base editing, prime editing, RNA targeting, and epigenetic modulation. While early clinical successes relied on ex vivo manipulation of patient-derived cells, recent advances in delivery chemistry and vector engineering are enabling direct in vivo editing across multiple organs. Here, we provide a comprehensive review of delivery modalities of CRISPR systems solely in vivo that underpin their therapeutic translation. We examine how anatomical, cellular, and immunological constraints shape organ-specific editing strategies in different organ systems and we highlight key preclinical and clinical milestones that define the current translational landscape. Across indications, delivery remains a critical determinant of efficacy, safety, and scalability, governing editor exposure, tissue selectivity, and risk of unintended genomic or epigenomic perturbation. This review, authored by members of the COST Action Genome Editing to treat Human Diseases (GenE-HumDi) Network, delineates the principles guiding in vivo genome and epigenome editing and outlines the remaining barriers to durable, tissue-selective, and broadly deployable CRISPR therapeutics.},
}

Humans
*Gene Editing/methods
Animals
*CRISPR-Cas Systems/genetics
Epigenome Editing
Genetic Therapy/methods
*Gene Transfer Techniques

@article {pmid41795918,
year = {2026},
author = {Liu, Y and Zhang, Y and Ding, H and Li, J and Gao, T and Wen, Z and Wang, Y and Wu, B},
title = {N-myristoyltransferase 1, a key gene for protein N-myristoylation, is dispensable for fertility in male mice.},
journal = {The Journal of reproduction and development},
volume = {72},
number = {4},
pages = {632-640},
doi = {10.1262/jrd.2025-064},
pmid = {41795918},
issn = {1348-4400},
mesh = {Animals ; Male ; *Acyltransferases/genetics/metabolism ; *Fertility/genetics/physiology ; *Spermatogenesis/genetics ; Mice ; Mice, Knockout ; Testis/metabolism ; Spermatozoa/metabolism ; Meiosis/genetics ; Infertility, Male/genetics ; CRISPR-Cas Systems ; },
abstract = {N-Myristoyltransferase 1 (NMT1), the predominant enzyme catalyzing myristoylation of proteins, is involved in various biological processes, including early embryonic development, immune responses, apoptosis, cellular homeostasis, tumorigenesis, and infection, with therapeutic potential in viral and parasitic infections as well as cancer. Despite the critical functions of NMT1, there have been no reports to date regarding its role in reproduction, especially in spermatogenesis. To investigate the function of NMT1 in this context, we utilized CRISPR/Cas9 technology to create a germ cell-specific Nmt1 knockout mice model for the first time. Surprisingly, male mice lacking NMT1 maintained fertility, exhibiting normal testicular structure and sperm morphology, with no significant differences in spermatogenic tubule structure or germ cell distribution compared to wild-type mice. Additionally, the Nmt1[f/f]; Stra8-Cre male mice showed no notable defects in meiosis. These findings suggest that NMT1 is not critical for spermatogenesis or male fertility in mice. However, further studies have shown that the compensatory role of NMT2 may play an unexpected role in maintaining myristoylation levels, which provides a new perspective for understanding the role of myristoylation in spermatogenesis.},
}

Animals
Male
*Acyltransferases/genetics/metabolism
*Fertility/genetics/physiology
*Spermatogenesis/genetics
Mice
Mice, Knockout
Testis/metabolism
Spermatozoa/metabolism
Meiosis/genetics
Infertility, Male/genetics
CRISPR-Cas Systems

@article {pmid41873043,
year = {2026},
author = {Lee, SJ and Lee, GS and Kim, J and Park, KH and Go, SR and Moon, JH and Woo, EJ},
title = {Mass spectrometry based identification of AMP-O-Tris generated by Thermococcus onnurineus Cas10.},
journal = {FEBS open bio},
volume = {16},
number = {8},
pages = {1593-1601},
pmid = {41873043},
issn = {2211-5463},
support = {Korean Government MSIP RS-2022-NR071772 RS-202//National Research Foundation of Korea/ ; KGM5382632 KGM1062612 KGM1322612//Korea Research Institute of Bioscience and Biotechnology/ ; CRC22024-500//National Research Council of Science and Technology/ ; },
mesh = {*Thermococcus/metabolism/genetics ; Tandem Mass Spectrometry/methods ; *Adenine Nucleotides/metabolism/chemistry ; CRISPR-Cas Systems/genetics ; Adenosine Triphosphate/metabolism ; Chromatography, High Pressure Liquid ; Bacterial Proteins/metabolism ; Adenosine Monophosphate/metabolism ; },
abstract = {Cas10, the catalytic core of type III CRISPR-Csm systems, synthesizes cyclic oligoadenylate (cOA) second messengers to activate downstream immune responses. Although Cas10 activity is regulated by complex assembly, the nucleophile selectivity and off-pathway reactivity of isolated Cas10 remain poorly understood. Here, using HPLC separation and subsequent tandem mass spectrometry (MS/MS) analysis, we identify and structurally characterize AMP-O-Tris as a noncanonical adenylylated product generated by isolated Thermococcus onnurineus Cas10. Our results reveal that purified Cas10 exhibits relaxed nucleophile selectivity, diverting ATP turnover into nonproductive adenylylation involving buffer-derived nucleophiles. This suggests that effector complex assembly constrains Cas10 reactivity to promote efficient cOA synthesis and suppress off-pathway chemistry. Furthermore, interception of reactive intermediates by buffer-derived nucleophiles may represent a potential chemical fail-safe that limits unintended signaling when Cas10 is uncoupled from the complex. Together, our study provides mechanistic insight into Cas10 regulation and informs the development of robust type III-based diagnostic platforms. Impact statement Our study reveals that Cas10 exhibits latent catalytic flexibility when isolated, identifying a noncanonical adenylation reaction. These findings demonstrate how complex assembly constrains enzymatic specificity to prevent aberrant signaling. This mechanistic insight is crucial for improving the fidelity and design of next-generation CRISPR-based diagnostic platforms.},
}

*Thermococcus/metabolism/genetics
Tandem Mass Spectrometry/methods
*Adenine Nucleotides/metabolism/chemistry
CRISPR-Cas Systems/genetics
Adenosine Triphosphate/metabolism
Chromatography, High Pressure Liquid
Bacterial Proteins/metabolism
Adenosine Monophosphate/metabolism

@article {pmid42089437,
year = {2026},
author = {Wei, S and Zhang, K and Deng, S and Chen, J and Huang, X and Guo, J and Wu, Y and Guo, Y and Liang, Z},
title = {Transposase-Assisted Donor Tethering Boosts Large-Fragment HDR in Plants.},
journal = {Advanced science (Weinheim, Baden-Wurttemberg, Germany)},
volume = {13},
number = {43},
pages = {e75565},
pmid = {42089437},
issn = {2198-3844},
support = {32170410//National Natural Science Foundation of China/ ; 62572289//National Natural Science Foundation of China/ ; 202403021221020//Fundamental Research Program of Shanxi Province/ ; 2023-006//Shanxi Scholarship Council of China/ ; 202203021224002//Natural Science Foundation of Shanxi Province for the Excellent Youth/ ; },
mesh = {*Transposases/genetics/metabolism ; *Recombinational DNA Repair/genetics ; CRISPR-Cas Systems/genetics ; Plants, Genetically Modified/genetics ; DNA Breaks, Double-Stranded ; },
abstract = {Precise insertion of large DNA fragments by homology-directed repair (HDR) remains inefficient and poorly reproducible in plants, largely due to limited donor availability at double-strand break sites. Here, we develop a transposase-assisted donor tethering strategy that improves the reliability of HDR-mediated large-fragment insertion. By fusing Cas9 to an integration-defective piggyBac variant that retains sequence-specific DNA-binding activity, donor templates are physically co-localized with Cas9-induced breaks. When combined with a transcription-coupled donor and a repair-pathway-biased Cas9 variant, this system enhances the frequency of accurate large-fragment insertions. Using this approach, we achieved efficient and precise kilobase-scale targeted gene insertions across multiple loci in both dicot and monocot species. These findings establish donor tethering as an effective strategy to improve plant HDR efficiency and provide a general framework for precise large-fragment genome insertion.},
}

*Transposases/genetics/metabolism
*Recombinational DNA Repair/genetics
CRISPR-Cas Systems/genetics
Plants, Genetically Modified/genetics
DNA Breaks, Double-Stranded

@article {pmid42309993,
year = {2026},
author = {Yu, W and Chen, J and Guo, J and Yu, F and Wang, G and Lin, J and Dai, X and Tan, X and Ma, P and Wu, L and Zhang, Y and Huang, S and Lan, P and Bian, Q and Huang, X and Wei, J and Cheng, T and Zheng, X and Qiao, Y},
title = {Guide RNA reprogramming facilitates minimized tracrRNA-dependent off-target and versatile CRISPR/Cas9 engineering.},
journal = {Nature communications},
volume = {17},
number = {1},
pages = {},
pmid = {42309993},
issn = {2041-1723},
mesh = {*CRISPR-Cas Systems/genetics ; Animals ; Humans ; *RNA, Guide, CRISPR-Cas Systems/genetics/metabolism ; Mice ; *Gene Editing/methods ; HEK293 Cells ; Machine Learning ; Transcriptome ; },
abstract = {While innovative, current CRISPR-Cas9 systems face safety concerns and practical hurdles, notably sequence-independent, noncanonical off-targeting. We demonstrate that the crRNA:tracrRNA duplex in guide RNAs (gRNA) is both splittable and reprogrammable. This property, however, enables endogenous RNAs with crRNA-like sequences to hijack any gRNAs, causing low-frequency yet pervasive tracrRNA-dependent off-target (TDO) effects. Using machine learning trained on high-throughput gRNA variant screens, we derive optimal gRNA-designing rules and engineer crRNA variants mismatched to the human/mouse transcriptomes, thereby minimizing TDO. By leveraging splittability and reprogrammability, we develop reprogrammable tracrRNAs for CRISPRa-based mRNA detection and redesign scaffolds to curb PAM-less Cas9-mediated "self-editing". We further create a separately expressed gRNA (segRNA) platform featuring split tracrRNAs and non-repetitive tandem crRNAs, enabling multiplexed editing of up to six genes and functional enhancer annotation in stem cells. Our findings uncover a previously overlooked off-target mechanism and offer versatile strategies to enhance the safety and utility of CRISPR systems.},
}

*CRISPR-Cas Systems/genetics
Animals
Humans
*RNA, Guide, CRISPR-Cas Systems/genetics/metabolism
Mice
*Gene Editing/methods
HEK293 Cells
Machine Learning
Transcriptome

@article {pmid42310027,
year = {2026},
author = {Busquets, O and Li, H and Syed, KM and Jerez, PA and Dunnack, J and Lo Bu, R and Verma, Y and Pangilinan, GR and Martin, A and Straub, J and Du, Y and Simon, VM and Poser, S and Bush, Z and Diaz, J and Sahagun, A and Gao, J and Hong, S and Hernandez, DG and Levine, KS and Pochet, N and Booth, EO and Blanchette, M and Bateup, HS and Rio, DC and Blauwendraat, C and Hockemeyer, D and Soldner, F},
title = {iSCORE-PD: an isogenic stem cell collection to research Parkinson's disease.},
journal = {Nature communications},
volume = {17},
number = {1},
pages = {},
pmid = {42310027},
issn = {2041-1723},
support = {1R56NS128015//U.S. Department of Health & Human Services | NIH | National Institute of Neurological Disorders and Stroke (NINDS)/ ; 1R01NS138402//U.S. Department of Health & Human Services | NIH | National Institute of Neurological Disorders and Stroke (NINDS)/ ; 1R01NS133140//U.S. Department of Health & Human Services | NIH | National Institute of Neurological Disorders and Stroke (NINDS)/ ; 5F31NS129265//U.S. Department of Health & Human Services | NIH | National Institute of Neurological Disorders and Stroke (NINDS)/ ; P30 CA013330/CA/NCI NIH HHS/United States ; P30 CA013330/CA/NCI NIH HHS/United States ; },
mesh = {Humans ; *Parkinson Disease/genetics/pathology ; Mutation ; Female ; Gene Editing ; alpha-Synuclein/genetics ; *Induced Pluripotent Stem Cells/metabolism ; Whole Genome Sequencing ; Cell Line ; CRISPR-Cas Systems ; *Pluripotent Stem Cells/metabolism ; Genetic Variation ; },
abstract = {Genome-edited human pluripotent stem cells (hPSCs) provide a powerful platform to study complex diseases such as Parkinson's disease (PD). Here, we describe iSCORE-PD, an isogenic collection of 65 genome-edited hPSC lines carrying disease-causing or high-risk variants in 11 PD-linked genes (SNCA, PRKN, PINK1, DJ1/PARK7, LRRK2, ATP13A2, FBXO7, DNAJC6, SYNJ1, VPS13C, and GBA1). All lines are derived from a well-characterized female hESC line and subjected to extensive quality control. Whole-genome sequencing reveals that genetic variation between lines, largely confined to non-coding regions, is minimal relative to inter-individual differences in patient-derived hiPSCs, with most variation arising from random mutations acquired during cell culture rather than genome-editing-induced off-target effects. Including multiple independently derived clones per mutation can control for this random genetic drift. Our systematic approach ensures high quality of this publicly available iSCORE-PD resource, highlights the advantages of prime editing over conventional CRISPR/Cas9 methods, and establishes best practices for generating disease-modeling hPSC collections.},
}

Humans
*Parkinson Disease/genetics/pathology
Mutation
Female
Gene Editing
alpha-Synuclein/genetics
*Induced Pluripotent Stem Cells/metabolism
Whole Genome Sequencing
Cell Line
CRISPR-Cas Systems
*Pluripotent Stem Cells/metabolism
Genetic Variation

@article {pmid42315520,
year = {2026},
author = {Chen, S and Hsiao, S and Xie, T and Chen, D and Chen, N and Jiang, J and Li, J and Wu, Y and Liao, J},
title = {Deep learning-guided engineering of SpuFz1 and rational miniaturization of ωRNA enables efficient genome editing.},
journal = {Nature communications},
volume = {17},
number = {1},
pages = {},
pmid = {42315520},
issn = {2041-1723},
mesh = {Humans ; Animals ; *Gene Editing/methods ; Mice ; *Deep Learning ; *Protein Engineering/methods ; RNA, Guide, CRISPR-Cas Systems/genetics ; *Endonucleases/genetics/metabolism ; HEK293 Cells ; Hematopoietic Stem Cells/metabolism ; CRISPR-Cas Systems ; *RNA/genetics ; Mutation ; },
abstract = {Advancing the performance of programmable genome editing nucleases remains a key challenge in expanding their research and therapeutic applications. Here, we introduce a scalable deep learning-guided protein engineering framework for improving nuclease activity without requiring experimental training data. As a demonstration, we apply this strategy to SpuFz1, a compact Fanzor nuclease of eukaryotic origin, identifying and validating beneficial mutations that produces a multi-mutant variant with an 11.6-fold increase in editing efficiency. In parallel, we use comparative sequence analysis to design and experimentally validate a 75-nt ultrashort ωRNA scaffold, reducing guide RNA length by 79% while maintaining activity. Integration of these optimized components yields enFanzor, a compact genome editing system that achieves editing efficiencies up to 81.9% in mammalian cells, with strong editing performance in both human hematopoietic stem and progenitor cells (HSPCs) and mouse embryos. The outperforming variant developed through this strategy also supports robust CBE and ABE activity. Notably, the shortened ωRNA not only improves nuclease editing specificity but also leads to a substantial increase in base editing efficiency. Together, this work demonstrates the power of combining AI-guided protein optimization with rational RNA design, and establishes a generalizable strategy for engineering next-generation genome editing tools.},
}

Humans
Animals
*Gene Editing/methods
Mice
*Deep Learning
*Protein Engineering/methods
RNA, Guide, CRISPR-Cas Systems/genetics
*Endonucleases/genetics/metabolism
HEK293 Cells
Hematopoietic Stem Cells/metabolism
CRISPR-Cas Systems
*RNA/genetics
Mutation

@article {pmid42333443,
year = {2026},
author = {},
title = {RNA-Triggered Chromatin Shredding Hits Cancer's Hardest Targets.},
journal = {Cancer discovery},
volume = {16},
number = {8},
pages = {OF1},
doi = {10.1158/2159-8290.CD-NW2026-0072},
pmid = {42333443},
issn = {2159-8290},
mesh = {Animals ; Humans ; *Chromatin/genetics/metabolism ; *Neoplasms/genetics/therapy ; Mice ; CRISPR-Cas Systems ; Mutation ; *RNA/genetics ; Tumor Suppressor Protein p53/genetics ; },
abstract = {RNA-triggered chromatin shredding may offer a new way to attack cancers driven by mutations that have resisted conventional drugs, two new studies show. In mouse models, upon activation by a target transcript, the CRISPR enzyme Cas12a2 can selectively eliminate tumor cells carrying mutations in TP53, MYC, and other hard-to-drug cancer genes.},
}

Animals
Humans
*Chromatin/genetics/metabolism
*Neoplasms/genetics/therapy
Mice
CRISPR-Cas Systems
Mutation
*RNA/genetics
Tumor Suppressor Protein p53/genetics

@article {pmid42511810,
year = {2026},
author = {Zaman, W and Ayaz, A},
title = {Beyond Permanent Genome Editing: Molecular Delivery Strategies for RNA Editing and Epigenome-Editing Therapeutics.},
journal = {International journal of molecular sciences},
volume = {27},
number = {14},
pages = {},
pmid = {42511810},
issn = {1422-0067},
mesh = {Humans ; *Epigenome Editing/methods ; *RNA Editing ; Animals ; *Gene Editing/methods ; CRISPR-Cas Systems ; RNA, Guide, CRISPR-Cas Systems/genetics ; *Gene Transfer Techniques ; },
abstract = {Reversible genetic medicines are emerging as controllable alternatives to permanent genome editing by enabling programmable modulation of RNA sequence, transcript abundance, chromatin state, and gene expression without irreversible genomic alteration. However, reversibility is not a single binary property: transient editor exposure, decay of the molecular effect, recovery of cellular function, and clinical capacity to stop, redose, or counteract treatment may diverge. This review therefore distinguishes mechanistic, functional, and clinical reversibility while examining targeted delivery systems for RNA-editing and epigenome-editing therapeutics. Key payloads include ADAR-recruiting oligonucleotides, CRISPR-Cas13 RNA editors, guide RNAs, chemically modified RNAs, editor-encoding mRNAs, dCas9 transcriptional regulators, DNA methylation editors, histone-modifying systems, and CRISPRoff-like platforms. We evaluate extracellular and intracellular delivery barriers, including nuclease degradation, immune recognition, renal clearance, liver uptake, cellular entry, endosomal escape, cytoplasmic release, nuclear localization, chromatin access, editing-window duration, off-target activity, immunogenicity, repeat-dosing feasibility, manufacturing, quality control, potency assays, and regulatory translation. Overall, delivery systems for reversible genetic medicines should be judged by tissue selectivity, functional editing, duration of action, reversibility after treatment withdrawal, safety, manufacturability, and clinical controllability.},
}

Humans
*Epigenome Editing/methods
*RNA Editing
Animals
*Gene Editing/methods
CRISPR-Cas Systems
RNA, Guide, CRISPR-Cas Systems/genetics
*Gene Transfer Techniques

@article {pmid42511812,
year = {2026},
author = {Liu, R and Cong, S and Gao, Y and Xu, J and Shi, X},
title = {Application of CRISPR-Cas9-Based Gene Editing Technology in Inherited Liver Diseases.},
journal = {International journal of molecular sciences},
volume = {27},
number = {14},
pages = {},
pmid = {42511812},
issn = {1422-0067},
support = {82300674//National Natural Science Foundation of China/ ; 2024BSL005//Liaoning Normal University Doctoral Research Start-up Project/ ; },
mesh = {Humans ; *Liver Diseases/genetics/therapy ; *Gene Editing/methods ; *CRISPR-Cas Systems ; Animals ; Genetic Therapy/methods ; },
abstract = {Inherited liver diseases are predominantly caused by monogenic mutations, and the vast majority of these conditions currently lack curative treatment options. Although liver transplantation may be used for patients with end-stage disease, it faces numerous challenges, including donor organ shortage, immune rejection, and the need for lifelong immunosuppression. In recent years, CRISPR-Cas9-based gene editing technology has advanced rapidly, offering transformative hope for the treatment of these diseases. This review systematically elucidates the working principles and technical advantages of the CRISPR-Cas9 system and its derived tools (base editing and prime editing), summarizes recent applications of these technologies in the treatment of hereditary liver diseases, and discusses the prospects and challenges of their clinical translation, aiming to provide a theoretical reference for future research in this field.},
}

Humans
*Liver Diseases/genetics/therapy
*Gene Editing/methods
*CRISPR-Cas Systems
Animals
Genetic Therapy/methods

@article {pmid42514096,
year = {2026},
author = {Wu, Y and Huang, X and Gu, Q and Li, Y and Zhou, Y and Sun, L and Wang, X},
title = {Characterization of CRISPR Loci and Antimicrobial Resistance in Foodborne Listeria monocytogenes Isolates.},
journal = {Microorganisms},
volume = {14},
number = {7},
pages = {},
pmid = {42514096},
issn = {2076-2607},
support = {2025KJ29//Nanjing Customs/ ; },
abstract = {Clustered regularly interspaced short palindromic repeats (CRISPR) are widespread in bacterial and archaeal genomes as an adaptive immune system against invading mobile genetic elements. This study investigated the distribution of CRISPR loci and their potential association with antimicrobial resistance (AMR) in 40 foodborne Listeria monocytogenes isolates. CRISPR analysis showed that 18 isolates harbored CRISPR Locus 1, five carried Locus 2, and five possessed both loci. Antimicrobial susceptibility testing against seven antimicrobial agents indicated that most isolates were highly susceptible to the tested agents. Specifically, all isolates were susceptible to gentamicin, ampicillin, penicillin, and tetracycline, whereas resistance was observed in a small subset of isolates: four were resistant to chloramphenicol, five to levofloxacin, and four to ciprofloxacin. Statistical analysis showed no statistically significant association between the presence of CRISPR loci and antimicrobial susceptibility phenotypes. These findings provide baseline information on CRISPR locus distribution and antimicrobial susceptibility profiles in foodborne L. monocytogenes isolates. Further studies based on larger isolate collections, whole-genome sequencing, and characterization of associated cas genes are needed to clarify the potential role of CRISPR-Cas systems in AMR evolution in this species.},
}

@article {pmid42514545,
year = {2026},
author = {Huiban, FA and Galović, V and Tripon, MR and Orlović, S and Tulcan, C and Camen, D},
title = {Nanotechnology-Enabled CRISPR Delivery: Emerging Opportunities in Agriculture and Forest Biotechnology.},
journal = {Plants (Basel, Switzerland)},
volume = {15},
number = {14},
pages = {},
pmid = {42514545},
issn = {2223-7747},
abstract = {Genome editing is one of the key technologies in contemporary plant biotechnology, which has been revolutionized using CRISPR/Cas systems that offer rapid, flexible, and precise options to improve agricultural characteristics, enhance stress tolerance, and accelerate breeding of crops and trees. Despite the significant benefits of CRISPR/Cas systems, their use is restricted by difficulties in genome-editing materials into plant cells. The conventional approaches include Agrobacterium-mediated transformation, particle bombardment and PEG-mediated transfection; these have contributed significantly to advancements in the field; however, dependent on specific plants and requiring tissue cultures, these methods lead to random transgene insertion and poor transformation efficiency. In addition, nanotechnology represents a novel method of delivering CRISPR cargos into plant cells using minimal invasiveness and potentially without DNA. This review provides a synopsis of the most employed CRISPR/Cas systems within plants, comparing the traditional delivery mechanisms and the various nanotechnological delivery vehicles, such as lipid nanoparticles, carbon nanotubes, DNA nanostructures, mesoporous silica nanoparticles, magnetically responsive nanoparticles and green nanomaterials. This review discusses the present challenges of delivery efficacy, biocompatibility, cargo integrity, and regulatory issues, and provides suggestions for future research directions regarding nanotechnology-assisted genome editing for precision breeding, sustainable agriculture, production of crops tolerant to climate conditions, and forest biotechnology.},
}

@article {pmid42514579,
year = {2026},
author = {Janeeshma, E and Das, S and Bouzroud, S and Sarraf, M and Akhtar, N and Mousavi, H and Ibrahimova, U and Fujita, M and Hasanuzzaman, M},
title = {Reactive Oxygen Species in Crop Plants: Production, Detoxification, Signaling, and Molecular Cross-Talk.},
journal = {Plants (Basel, Switzerland)},
volume = {15},
number = {14},
pages = {},
pmid = {42514579},
issn = {2223-7747},
abstract = {Although excess generation of reactive oxygen species (ROS) is harmful for plants, at mild concentrations, they are able to positively regulate the metabolic pathway and signaling cascades of the cell. The mechanisms functional under the ROS-induced stress tolerance need to be decoded for the development of new abiotic stress-tolerant crop varieties with a clear vision. This study details ROS generation, roles of enzymatic antioxidants as well as non-enzymatic antioxidants, and ROS-induced cellular events. Emphasis is given to crucial topics such as cyclin-dependent kinases and mitogen-activated protein kinases signaling mechanisms, calcium-mediated cellular cross-talk, and activation or inactivation of various transcription factors. Introduction of different genetic and molecular approaches to manipulate the ROS pathway helps to find out novel recombinant plant varieties. Selection of specific biotechnological tools, appropriate omics analysis and implementation of CRISPR/Cas 9 in crop plants assist the designing of mutants. This review highlights the beneficial roles of oxidative stress, spotlighting the molecular mechanisms that support the physiological, morphological and biochemical modifications of plant cells.},
}

@article {pmid42516529,
year = {2026},
author = {Nikravan, M and Heidari, H and Khoshnood, S and Ghafourian, S and Kazemian, H},
title = {Molecular Profiling of CRISPR-Cas System, Virulence Traits, and Antimicrobial Resistance in Enterococcus faecalis Clinical Isolates.},
journal = {International journal of microbiology},
volume = {2026},
number = {},
pages = {5903066},
pmid = {42516529},
issn = {1687-918X},
abstract = {INTRODUCTION: Enterococcus faecalis is responsible for life-threatening enterococcal infections. This study is aimed at investigating virulence factors, antimicrobial resistance patterns, and molecular characteristics of clinical E. faecalis isolates.

MATERIALS AND METHODS: A total of 42 E. faecalis isolates were collected. Antimicrobial resistance and the minimum inhibitory concentrations (MICs) of vancomycin and gentamicin were determined using standard microbroth dilution method. The presence of virulence, antibiotic resistance, and CRISPR-Cas genes was investigated by polymerase chain reaction (PCR). Biofilm formation was also assessed, and the genetic diversity of the isolates was analyzed using enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR).

RESULTS: The highest resistance rates were observed for gentamicin and ampicillin, and all isolates were high-level gentamicin resistant (HLGR) (MICs ≥ 500 μg/mL). Vancomycin resistance was detected in 15 isolates (35.7%) (MICs ≥ 32 μg/mL). Thirty-nine isolates (92.8%) were biofilm producers. The efaA gene was the most frequently detected (95.2%), followed by esp (69%), gelE (66.6%), ace (66.6%), and asa1 (57.1%). The aac(6  ['])-Ie-aph(2  [″])-Ia gene was detected in all isolates (100%), followed by ermB (78.6%), ermA (38.1%), ermC (23.8%), and vanA (19%). CRISPR3 was the most frequently detected locus (80.95%), followed by CRISPR1 (50%), CRISPR1-cas csn1 (23.8%), and CRISPR2 (9.5%). The CRISPR3-cas csn1 gene was not detected in any isolate. High heterogeneity was observed among the isolates, with 35 different ERIC types identified.

CONCLUSION: This study demonstrated notable resistance traits and high genetic diversity among clinical E. faecalis isolates, but no statistically significant association was found between CRISPR-Cas genes and phenotypic or genotypic resistance.},
}

@article {pmid42517263,
year = {2026},
author = {Hu, X and Xu, P and Shen, M and Chai, X and Tan, J and Sun, X and Yang, L and Wang, Q and Tan, F},
title = {A CRISPR-Cas12a and Quantum-Dot Lateral-Flow Assay for Rapid Species-Level Detection of Trichophyton rubrum.},
journal = {Mycoses},
volume = {69},
number = {7},
pages = {e70208},
pmid = {42517263},
issn = {1439-0507},
support = {22Y11905700//Program of the Science and Technology Commission of Shanghai Municipality "Science and Technology Innovation Action Plan" Medical Innovation Research Special Project/ ; SHDC12026111//Shanghai Shen Kang Clinical Technology Promotion Program/ ; YXX2024-KF01-04//The Center for Basic Research and Innovation of Medicine and Pharmacy (MOE)/ ; 2026ms03//Xi'an Municipal Health Commission General Research Project/ ; },
mesh = {Humans ; Sensitivity and Specificity ; *CRISPR-Cas Systems ; *Quantum Dots ; Rapid Diagnostic Tests ; *Arthrodermataceae/isolation & purification/genetics ; *Tinea/diagnosis/microbiology ; *Trichophyton/isolation & purification/genetics ; *Molecular Diagnostic Techniques/methods ; Real-Time Polymerase Chain Reaction ; DNA, Fungal/genetics ; },
abstract = {BACKGROUND: Accurate species-level diagnosis of dermatophytes, particularly Trichophyton rubrum, is important for guiding targeted antifungal therapy and improving the management of recurrent or atypical superficial mycoses. However, routine microscopy has only moderate sensitivity and cannot resolve fungal species; fungal culture is time-consuming, and real-time quantitative PCR (qPCR) requires costly instrumentation.

OBJECTIVES: To develop and clinically evaluate CRI-RUB, a CRISPR-Cas12a-based assay integrating recombinase-aided amplification (RAA) and a quantum-dot (QD) fluorescent lateral-flow strip for rapid species-level detection of T. rubrum.

PATIENTS/METHODS: Analytical sensitivity and specificity were assessed using plasmid standards and a panel of common cutaneous fungi. A total of 140 clinical specimens were tested by both CRI-RUB and TaqMan qPCR. Discrepant or grey-zone qPCR results (Ct 33-34), together with a subset of concordant cases, were further arbitrated by internal transcribed spacer (ITS) sequencing to establish a composite reference standard.

RESULTS: CRI-RUB achieved a visual limit of detection of 2.3 × 10[1] copies/μL and showed no cross-reactivity with other common cutaneous fungi. Among the 140 clinical specimens, CRI-RUB showed a sensitivity of 96.1% (73/76), specificity of 100.0% (64/64), positive predictive value of 100.0% (73/73), negative predictive value of 95.5% (64/67) and Cohen's κ of 0.96 compared with the composite reference standard, indicating excellent concordance. The complete workflow required approximately 85 min.

CONCLUSIONS: CRI-RUB provides a rapid, sensitive, and specific approach for species-level detection of T. rubrum. By combining CRISPR-Cas12a detection with QD-based lateral-flow readout, this assay shows potential for near-patient or point-of-care dermatophyte diagnosis.},
}

Humans
Sensitivity and Specificity
*CRISPR-Cas Systems
*Quantum Dots
Rapid Diagnostic Tests
*Arthrodermataceae/isolation & purification/genetics
*Tinea/diagnosis/microbiology
*Trichophyton/isolation & purification/genetics
*Molecular Diagnostic Techniques/methods
Real-Time Polymerase Chain Reaction
DNA, Fungal/genetics

@article {pmid42510959,
year = {2026},
author = {Gubaidullin, N and Ospankulova, G and Gajimuradova, A and Syzdykova, A and Zhumalin, A and Dairova, K and Konysbayeva, D and Gorbulya, V and Makangali, K},
title = {ROS-Centered Transcriptomic Regulatory Networks Linking Salinity Stress, Antioxidant Defense and Processability Traits in Salicornia spp.},
journal = {Current issues in molecular biology},
volume = {48},
number = {7},
pages = {},
pmid = {42510959},
issn = {1467-3045},
support = {BR22883587//the ministry of agriculture of Kazakhstan/ ; },
abstract = {Salinity stress affects not only the survival and productivity of halophytic plants, but also the composition, structure and processability of their biomass. In Salicornia spp., salt-induced regulation of ion transport, osmotic adjustment, reactive oxygen species signaling, antioxidant defense, and cell wall remodeling can directly influence residual salinity, water retention, texture, extractability, drying behavior, and oxidative stability of plant biomass. However, most existing transcriptomic studies of Salicornia and related halophytes have focused mainly on salt tolerance mechanisms, whereas the connection between stress-regulated molecular networks and processing-related biomass traits remains insufficiently systematized. This review addresses this gap by proposing a mechanistic framework that links salinity perception, ROS-mediated signaling, ABA and MAPK pathways, antioxidant gene families, transcription factor networks and processing-oriented quality traits. Special attention is given to enzymatic antioxidant systems, including SOD, CAT, APX, POD and components of the ascorbate-glutathione cycle, as well as to non-enzymatic defense mechanisms involving ascorbate, glutathione, phenolic compounds, carotenoids, proline and glycine betaine. The review also discusses the regulatory roles of WRKY, DREB/CBF, NAC, bZIP and MYB transcription factor families as molecular control points connecting salinity stress responses with downstream metabolic and structural traits. Network-based approaches, including WGCNA, pathway signatures and transcript panels, are considered more informative than single-gene markers for predicting complex quality traits in Salicornia biomass. In addition, recent genomic and computational strategies, including CRISPR/Cas-mediated functional validation, GWAS, genomic selection, multi-omics integration and AI-assisted modeling, are discussed as emerging tools for candidate-gene prioritization and predictive assessment of stress-dependent biomass quality. Overall, this review shifts the interpretation of Salicornia transcriptomics from a descriptive salt-tolerance model toward a mechanistic and application-oriented framework for improving halophytic raw materials for food, feed and bioprocessing applications.},
}

@article {pmid42511636,
year = {2026},
author = {Zhan, Y and Luo, T and Sun, Y},
title = {Efficient Gene Editing in Fish Primary Germline Stem Cells.},
journal = {International journal of molecular sciences},
volume = {27},
number = {14},
pages = {},
pmid = {42511636},
issn = {1422-0067},
support = {2022YFD2400101//The National Key R&D Program of China/ ; },
mesh = {Animals ; *Gene Editing/methods ; CRISPR-Cas Systems ; Transfection/methods ; Oryzias/genetics ; *Stem Cells/metabolism/cytology ; *Germ Cells/metabolism/cytology ; *Eels/genetics ; Carps/genetics ; },
abstract = {Genome editing by the CRISPR/Cas9 system is widely used for production of gene-modified animals, including fish. However, efficient gene editing in fish cultured cells, in particular germline stem cells (GSCs), is challenging, likely due to the difficulty in transfecting these cells. The ricefield eel (Monopterus albus), a sequential hermaphroditic species, is a freshwater fish of significant economic value in China. In this work, we report a simple method that can achieve high gene editing efficiency in primary GSCs of fish species, including ricefield eel. High transfection efficiency (50-95%) is achieved in fish GSCs by using a microchannel-based cell transfection system. Gene editing efficiency of up to 60% in primary ricefield eel GSCs is achieved using an integrated CRISPR/Cas9 vector strategy. High gene editing efficiency is also achieved in gibel carp (Carassius gibelio) GSCs and the medaka spermatogonial stem cell line SG3, demonstrating the general applicability of our method. Our data suggest that efficient transfection is key to high gene editing efficiency in fish cultured GSCs. This study establishes an efficient and reliable gene editing system for fish GSCs, which may facilitate the creation of new germplasm of genetically difficult-to-breed fish by combining GSC transplantation with gene editing techniques.},
}

Animals
*Gene Editing/methods
CRISPR-Cas Systems
Transfection/methods
Oryzias/genetics
*Stem Cells/metabolism/cytology
*Germ Cells/metabolism/cytology
*Eels/genetics
Carps/genetics

@article {pmid42480477,
year = {2026},
author = {Toishigawa, K and Magoori, K and Sato, H and Edahiro, T and Ureshino, H and Shindo, T and Suzuki, R and Sakuma, T and Yamamoto, T and Okada, M and Ichinohe, T},
title = {Platinum TALEN-mediated nonviral gene editing facilitates clinical-scale production of cancer antigen-reactive T cells.},
journal = {Cytotherapy},
volume = {28},
number = {9},
pages = {102911},
doi = {10.1016/j.jcyt.2026.102911},
pmid = {42480477},
issn = {1477-2566},
mesh = {Humans ; *Transcription Activator-Like Effector Nucleases/genetics/metabolism ; *Gene Editing/methods ; *Antigens, Neoplasm/immunology/genetics ; *T-Lymphocytes/immunology ; *Receptors, Antigen, T-Cell/genetics ; CRISPR-Cas Systems/genetics ; *Neoplasms/immunology/therapy ; },
abstract = {BACKGROUND AIMS: Recently, target-genome editing has emerged as a next-generation tool for the clinical development of designed cellular products. Although Clustered Regularly Interspaced Short Palindromic Repeats-Cas9 (CRISPR-Cas9) is the most frequently adopted nuclease for therapeutic genome editing, its widespread use is still hampered by potential off-target effects and high patent royalties. Platinum transcription activator-like effector nuclease (TALEN) is a modified TALEN that harbors non-repeat-variable di-residue (non-RVD) variations and confers higher efficiency than conventional TALENs lacking non-RVD variations.

METHODS: In this study, using Platinum TALEN targeting T-cell receptor (TCR) gene loci and a single-stranded DNA homology-directed repair (HDR) template, we aimed to produce TCR-replaced human T cells reprogrammed to recognize a cancer antigen and kill cancer cells.

RESULTS: This system can reproducibly produce TCR-engineered T cells from ∼50 mL of peripheral blood on a clinical scale. The resulting genome-edited T cells retain naïve/naïve-like and memory phenotype cells in both CD4+ and CD8+ fractions and exhibit efficient cytolytic activity against cancer cell lines in vitro.

CONCLUSION: In conclusion, Platinum TALEN-mediated nonviral genome editing facilitates the replacement of the endogenous TCR with a desired TCR and can be applied to the clinical manufacturing of therapeutic T-cell products.},
}

Humans
*Transcription Activator-Like Effector Nucleases/genetics/metabolism
*Gene Editing/methods
*Antigens, Neoplasm/immunology/genetics
*T-Lymphocytes/immunology
*Receptors, Antigen, T-Cell/genetics
CRISPR-Cas Systems/genetics
*Neoplasms/immunology/therapy

@article {pmid42510814,
year = {2026},
author = {Zhang, X and Lu, C and Hu, Z and Geng, X and Li, G and Cai, J},
title = {Complete Genome Analysis of Pectobacterium brasiliense BS1113, a Causal Agent of Cigar Tobacco Soft Rot, with Phenotypic Characterization of Virulence and Copper Tolerance.},
journal = {Genes},
volume = {17},
number = {7},
pages = {},
pmid = {42510814},
issn = {2073-4425},
support = {This research was funded by the Major Research Project of Fuyang Normal University (2025FSKJ31), the Doctoral Talent Introduction Project of Fuyang Normal University (2020KYQD0031), the Major Project of Anhui Huatuo Academy of Traditional Chinese Medicine//Fuyang Normal University/ ; },
mesh = {*Copper/metabolism/toxicity ; *Plant Diseases/microbiology/genetics ; *Nicotiana/microbiology ; Virulence/genetics ; *Genome, Bacterial ; Phenotype ; Whole Genome Sequencing ; },
abstract = {Background:Pectobacterium brasiliense-mediated soft rot severely threatens the production of diverse cash crops worldwide and brings severe yield reduction risks. A virulent strain BS1113 was separated from diseased cigar tobacco plants collected in Yunnan, yet its virulence regulatory genes and copper resistance-related genetic background have not been fully analyzed so far. This study aims to decipher the genomic features of BS1113 and clarify its pathogenic and copper-tolerant characteristics via whole-genome sequencing, comparative genomics and indoor phenotype verification. Methods: Hybrid sequencing strategies combining Illumina short reads and PacBio long reads were adopted to obtain the complete circular genome sequence of strain BS1113. Subsequent comparative genomic analysis and multiple phenotypic identification experiments were conducted to characterize its genetic architecture and physiological traits. Results: Genome assembly results showed that the circular chromosome of BS1113 spans 4,916,962 bp with a GC content of 51.96%, which encodes a total of 4369 functional protein-coding genes. Genomic comparison revealed that BS1113 completely lacks the T3SS gene cluster, while it conserves intact T2SS, T6SS and I-F CRISPR-Cas systems; the chromosomal copper resistance operon copRSAB was also detected in this isolate. Pathogenicity tests validated that BS1113 satisfies all criteria of Koch's postulates on cigar tobacco hosts. In addition, BS1113 displayed prominent tolerance against eight mainstream copper bactericides widely used for tobacco disease management. Conclusions: This research generates the first complete high-quality genome of P. brasiliense isolated from cigar tobacco hosts. The genomic data explain the infection mechanism of this pathogen independent of intact T3SS, and also reveal the genetic basis supporting its persistent survival under long-term copper fungicide pressure in field cultivation environments.},
}

*Copper/metabolism/toxicity
*Plant Diseases/microbiology/genetics
*Nicotiana/microbiology
Virulence/genetics
*Genome, Bacterial
Phenotype
Whole Genome Sequencing

@article {pmid42510829,
year = {2026},
author = {Yoshino, H and Fukuda, I and Enokida, H and Seki, N and Goto, Y},
title = {MEK1 as a Synthetic Lethal Target with Cabozantinib in Renal Cell Carcinoma: Insights from CRISPR/Cas9 Screening.},
journal = {Genes},
volume = {17},
number = {7},
pages = {},
pmid = {42510829},
issn = {2073-4425},
support = {24K12458//Japan Society for the Promotion of Science/ ; 24K11347//Japan Society for the Promotion of Science/ ; 24K12641//Japan Society for the Promotion of Science/ ; },
mesh = {Humans ; *Pyridines/pharmacology ; *Carcinoma, Renal Cell/genetics/drug therapy/pathology ; *Anilides/pharmacology ; Animals ; *Kidney Neoplasms/genetics/drug therapy/pathology ; *MAP Kinase Kinase 1/genetics/antagonists & inhibitors ; CRISPR-Cas Systems/genetics ; Mice ; Cell Line, Tumor ; *Synthetic Lethal Mutations ; Xenograft Model Antitumor Assays ; Protein Kinase Inhibitors/pharmacology ; Cell Proliferation/drug effects ; Cell Survival/drug effects ; },
abstract = {Background/Objectives: Cabozantinib is a tyrosine kinase inhibitor that primarily targets MET. It has become an important drug in the treatment of renal cell carcinoma (RCC); however, many patients do not respond to cabozantinib treatment and there is no effective next-line therapy. In this study, we identified molecular-targeted drugs that exhibit synergistic effects with cabozantinib using CRISPR/Cas9 screening. Methods: A kinome-wide synthetic lethal CRISPR/Cas9 screen was used to identify target molecules using 786-o RCC cells. A library was generated, and treatment with vehicle or cabozantinib was carried out, followed by next-generation sequencing to identify candidate genes. A combination index based on the Chou-Talalay method was used to evaluate the synergistic effect of cabozantinib through cell viability assays. Xenograft assays were conducted to determine the effect in vivo. Results: CRISPR/Cas9-based screening revealed four genes (MEK1, DCLK1, DYRK3, and FGFR1) that were candidates for synthetic lethality by cabozantinib in RCC cells. We focused on MEK1 because the MEK1 inhibitor cobimetinib has been approved for melanoma treatment. In a cell proliferation assay using 786-o and A498 RCC cells, the combination of cobimetinib and cabozantinib exhibited a synergistic effect. A xenograft assay also revealed a significant synergistic effect of cobimetinib and cabozantinib. Conclusions: CRISPR/Cas9 screening identified MEK1 as a candidate for a synthetic lethal target with cabozantinib in RCC. The combined inhibition of MET/VEGFR and MEK1 suppressed compensatory MAPK reactivation and downregulated the PI3K-Akt pathway, including the survival-associated genes PPP2R3B and ATF6B, and produced significant tumor growth suppression in vivo. These findings highlight the potential of cabozantinib plus cobimetinib, an already-FDA-approved MEK inhibitor, as a readily translatable combination strategy to overcome cabozantinib resistance in RCC.},
}

Humans
*Pyridines/pharmacology
*Carcinoma, Renal Cell/genetics/drug therapy/pathology
*Anilides/pharmacology
Animals
*Kidney Neoplasms/genetics/drug therapy/pathology
*MAP Kinase Kinase 1/genetics/antagonists & inhibitors
CRISPR-Cas Systems/genetics
Mice
Cell Line, Tumor
*Synthetic Lethal Mutations
Xenograft Model Antitumor Assays
Protein Kinase Inhibitors/pharmacology
Cell Proliferation/drug effects
Cell Survival/drug effects

@article {pmid42510848,
year = {2026},
author = {Zafar, MA and Harling, LC and Li, Y and Celik, NB and Mukherjee, SK and Rizzo, J and Prendergast, A and Elefteriades, JA},
title = {A Preliminary Zebrafish Model of ACTA2 Deficiency Reveals Increased Larval Phenotype Burden and Suggests Reduced Adult Mutant Survival.},
journal = {Genes},
volume = {17},
number = {7},
pages = {},
pmid = {42510848},
issn = {2073-4425},
support = {N/A//Masone Family/ ; Walter-Benjamin Scholarship//German Research Foundation/ ; },
mesh = {Animals ; *Zebrafish/genetics ; *Actins/genetics/deficiency ; Phenotype ; Larva/genetics ; Mutation ; *Zebrafish Proteins/genetics/deficiency ; Disease Models, Animal ; Genotype ; CRISPR-Cas Systems ; },
abstract = {BACKGROUND: ACTA2 encodes smooth muscle alpha-actin and is one of the most common genetic causes of inherited non-syndromic thoracic aortic aneurysm and dissection. Although murine models have provided important mechanistic insight, complementary vertebrate systems may enable more rapid in vivo phenotyping and future therapeutic screening.

METHODS: Utilizing a CRISPR/Cas9-based approach, we developed zebrafish acta2 mutant models and assessed phenotypes at larval and adult stages. At 3 days post-fertilization, larvae were screened by brightfield microscopy for edema, axis defects, hemorrhage, and a composite endpoint (any phenotype) under basal conditions and after exposure to 0.2 mM epinephrine. Phenotype occurrence was summarized as group-specific proportions across a 2 × 2 design defined by genotype (wild-type vs. acta2-deficient mutant) and treatment (DMSO control vs. epinephrine), with prespecified pairwise comparisons using Pearson's chi-squared tests. Pooled fish-level logistic regression models were also fit for each endpoint. Adult follow-up was performed by genotyping all available acta2 fish at the facility and tracking staggered tank-level cohorts longitudinally.

RESULTS: Initial gross morphologic assessment did not reveal overt external phenotypic differences between heterozygous or homozygous acta2 mutant larvae and wild-type controls. In larval analyses, the mutant genotype was associated with a greater burden of adverse phenotypes than treatment exposure. For the composite endpoint, mutant larvae demonstrated higher proportions than normal larvae under both control (28.3% vs. 17.4%, p = 0.007) and epinephrine conditions (26.9% vs. 17.4%, p = 0.023), whereas epinephrine did not significantly alter composite phenotype frequency within either genotype. Similar genotype-associated trends were observed for axis defects and hemorrhage. In pooled logistic regression, acta2 deficiency was associated with increased odds of axis defects (OR 2.64, 95% CI 1.48-4.89, p = 0.001), hemorrhage (OR 2.40, 95% CI 1.09-5.68, p = 0.036), and the composite endpoint (OR 1.88, 95% CI 1.19-3.00, p = 0.008), whereas epinephrine exposure did not demonstrate a consistent independent effect across endpoints. Adult staggered follow-up showed greater attrition in homozygous mutant cohorts than in wild-type or heterozygous cohorts, with aggregated losses of 24%, 12%, and 3%, respectively.

CONCLUSIONS: This preliminary zebrafish acta2 model demonstrated that acta2 deficiency is associated with increased adverse larval phenotype burden and suggested reduced long-term persistence of homozygous mutant fish in adulthood. The strongest signal in the current study was genotype-associated phenotype burden rather than a robust epinephrine-dependent effect. These findings support the feasibility of zebrafish-based ACTA2 phenotyping while highlighting the need for more specific vascular endpoints, refined longitudinal follow-up, and future variant-specific modeling.},
}

Animals
*Zebrafish/genetics
*Actins/genetics/deficiency
Phenotype
Larva/genetics
Mutation
*Zebrafish Proteins/genetics/deficiency
Disease Models, Animal
Genotype
CRISPR-Cas Systems

@article {pmid42259495,
year = {2026},
author = {Chandrapalan, T and Kwong, RWM},
title = {DMT1 regulates systemic trace metal handling and developmental outcomes: Insights from targeted mutagenesis in zebrafish.},
journal = {Comparative biochemistry and physiology. Toxicology & pharmacology : CBP},
volume = {308},
number = {},
pages = {110597},
doi = {10.1016/j.cbpc.2026.110597},
pmid = {42259495},
issn = {1532-0456},
mesh = {Animals ; *Zebrafish/genetics/metabolism/growth & development ; *Cation Transport Proteins/genetics/metabolism ; *Zebrafish Proteins/genetics/metabolism ; Solute Carrier Family 11, Member 2 ; *Trace Elements/metabolism ; Mutagenesis ; Gene Expression Regulation, Developmental ; CRISPR-Cas Systems ; Gene Knockout Techniques ; },
abstract = {Divalent metal transporter 1 (DMT1) is thought to be the primary route for non-heme iron absorption in vertebrates, but its systemic role remains poorly understood. Using CRISPR-Cas9 gene editing, we generated a DMT1 knockout (dmt1[-/-]; KO) zebrafish mutant line to examine the developmental and physiological consequences of DMT1 loss. Phenotypic and hematological assessments were performed alongside measurements of whole-body and tissue-specific metal concentrations. Further, to identify potential compensatory pathways during DMT1 loss, the expression profile of candidate metal transporters or ion channels (hcp1, zip4, zip8, zip14, and ecac) was quantified using droplet digital PCR (ddPCR). DMT1 KO larvae exhibited delayed development, anemia, and broad disruption in multiple trace metals (iron, zinc, manganese, cobalt, and selenium). Gene expression analysis during early development revealed higher hcp1 mRNA abundance in the mutant, suggesting a possible compensatory response to maintain metal homeostasis during DMT1 loss. Although viable to adulthood, the mutants had persisting iron dysregulation and red blood cell abnormalities. This study provides the first in vivo evidence of the physiological role of DMT1 in multi-metal balance in fish and offers new insight into compensatory mechanisms underlying DMT1 deficiency.},
}

Animals
*Zebrafish/genetics/metabolism/growth & development
*Cation Transport Proteins/genetics/metabolism
*Zebrafish Proteins/genetics/metabolism
Solute Carrier Family 11, Member 2
*Trace Elements/metabolism
Mutagenesis
Gene Expression Regulation, Developmental
CRISPR-Cas Systems
Gene Knockout Techniques

@article {pmid42288133,
year = {2026},
author = {Abrahms, ZN and Majdi, M and Madsen, SM and Morton, CJ and Sen, AK and Fried, NB and Sawyer, LE and Cegielski, ER and Spezzano, SJ and Jones, JA},
title = {Genome-based optimization of psilocybin and N,N-dimethyltryptamine biosynthetic pathways in E. coli using CRISPR-associated transposases.},
journal = {Metabolic engineering},
volume = {97},
number = {},
pages = {102490},
doi = {10.1016/j.ymben.2026.102490},
pmid = {42288133},
issn = {1096-7184},
mesh = {*Escherichia coli/genetics/metabolism ; *Psilocybin/biosynthesis/genetics ; *Metabolic Engineering/methods ; *Genome, Bacterial ; *CRISPR-Cas Systems ; *Biosynthetic Pathways/genetics ; },
abstract = {Stable, high-level biosynthesis of complex natural products requires precise control of heterologous pathway expression, yet transcriptional architectures optimized on plasmids often fail when transferred to the chromosome. Here, we present ePathIntegrate, a genome-centric pathway engineering strategy that leverages CRISPR-associated transposases (CASTs) to integrate and rebalance multigene metabolic pathways in Escherichia coli. Direct genomic transfer of plasmid-optimized psilocybin and N,N-dimethyltryptamine (DMT) pathways resulted in a loss of productivity, driven by context-dependent promoter behavior. To address this, we developed and characterized a library of mutant T7 promoters that restore mid-range transcriptional control on the genome. Applying ePathIntegrate enabled re-optimization of both pathways, yielding genome-encoded strains that achieve 1.88 g/L psilocybin and 1.62 g/L DMT in fed-batch bioreactors. Whole-genome sequencing of CAST-mediated strains further revealed (i) precise on-target integration, (ii) some off-target pathway integrations, and (iii) small mutations in a subset of strains, highlighting both the power and limitations of CAST-mediated strain engineering.},
}

*Escherichia coli/genetics/metabolism
*Psilocybin/biosynthesis/genetics
*Metabolic Engineering/methods
*Genome, Bacterial
*CRISPR-Cas Systems
*Biosynthetic Pathways/genetics

@article {pmid42297823,
year = {2026},
author = {Adler, KM and Xu, H and Gladstein, AC and Irizarry-Negron, VM and Robertson, MR and Doerig, KR and Petrov, DA and Winslow, MM and Feldser, DM},
title = {Tumor suppressor genotype influences the extent and mode of immunosurveillance in lung cancer.},
journal = {Nature communications},
volume = {17},
number = {1},
pages = {},
pmid = {42297823},
issn = {2041-1723},
support = {R01-CA262619-04//U.S. Department of Health & Human Services | NIH | National Cancer Institute (NCI)/ ; R01-CA-279698-01)//U.S. Department of Health & Human Services | NIH | National Cancer Institute (NCI)/ ; },
mesh = {Animals ; *Lung Neoplasms/genetics/immunology/pathology ; Genotype ; *Genes, Tumor Suppressor ; Humans ; *Immunologic Surveillance/genetics ; Immunoediting, Cancer ; Antigens, Neoplasm/immunology/genetics ; Proto-Oncogene Proteins p21(ras)/genetics ; Mice ; CRISPR-Cas Systems ; Cell Line, Tumor ; },
abstract = {The impact of cancer driving mutations on immunosurveillance throughout tumor development remains poorly understood. To better understand the contribution of tumor genotype to immunosurveillance, we generated and validated lentiviral-based vectors that create increasingly immunogenic neoantigens. This vector system is compatible with autochthonous Cre-regulated cancer models, CRISPR/Cas9-mediated somatic genome editing, and tumor barcoding. Here, we show that in the context of oncogenic KRAS-driven lung cancer and strong neoantigen expression, tumor suppressor genotype dictates the degree of immune cell recruitment, positive selection of tumors with neoantigen silencing, and tumor outgrowth. By quantifying the impact of 11 commonly inactivated tumor suppressor genes on tumor growth across neoantigenic contexts, we show that the growth-promoting effects of tumor suppressor gene inactivation correlate with increasing sensitivity to immunosurveillance. Importantly, some genotypes also dramatically changed sensitivity to immunosurveillance independently of their growth-promoting effects. We propose a model of immunoediting in which tumor suppressor gene inactivation works in tandem with neoantigen expression to shape tumor immunosurveillance and immunoediting such that the same neoantigens uniquely modulate tumor immunoediting depending on the genetic context.},
}

Animals
*Lung Neoplasms/genetics/immunology/pathology
Genotype
*Genes, Tumor Suppressor
Humans
*Immunologic Surveillance/genetics
Immunoediting, Cancer
Antigens, Neoplasm/immunology/genetics
Proto-Oncogene Proteins p21(ras)/genetics
Mice
CRISPR-Cas Systems
Cell Line, Tumor

@article {pmid42479086,
year = {2026},
author = {Ma, L and Zhao, J and Chen, Y and Wang, Z and Yan, B and Liu, Z and Ma, N},
title = {RI-augmented TdT-CRISPR fluorescent biosensor for sensitive quantification of DNA breakage in CT-irradiated human sperm.},
journal = {Analytical methods : advancing methods and applications},
volume = {18},
number = {29},
pages = {6090-6094},
doi = {10.1039/d6ay01260d},
pmid = {42479086},
issn = {1759-9679},
mesh = {Humans ; *Spermatozoa/radiation effects/metabolism ; *Biosensing Techniques/methods ; Male ; *DNA Damage ; *DNA ; *CRISPR-Cas Systems ; Limit of Detection ; },
abstract = {Featuring a linear range of 0.001-0.4 nM and a limit of detection of 0.12 pM, the RI-enhanced TdT-CRISPR sensor precisely detects radiation-induced trace DNA damage in sperm, outperforming the conventional DFI method.},
}

Humans
*Spermatozoa/radiation effects/metabolism
*Biosensing Techniques/methods
Male
*DNA Damage
*DNA
*CRISPR-Cas Systems
Limit of Detection

@article {pmid42482463,
year = {2026},
author = {Yu, J and Wang, J and Yang, Y and Sun, L and Hu, X and Nie, T and Yang, X and Wang, X and Li, C and You, X},
title = {Multifaceted effects of galU deletion on phenotype and virulence of Pseudomonas aeruginosa in vitro and in vivo.},
journal = {Virulence},
volume = {17},
number = {1},
pages = {2707803},
doi = {10.1080/21505594.2026.2707803},
pmid = {42482463},
issn = {2150-5608},
mesh = {*Pseudomonas aeruginosa/pathogenicity/genetics/drug effects ; Virulence ; Animals ; Pseudomonas Infections/microbiology ; Anti-Bacterial Agents/pharmacology ; *Gene Deletion ; *Bacterial Proteins/genetics/metabolism ; Phenotype ; O Antigens/genetics/biosynthesis ; Polymyxins/pharmacology ; Quorum Sensing ; Microbial Sensitivity Tests ; Mice ; CRISPR-Cas Systems ; },
abstract = {Pseudomonas aeruginosa is a widespread Gram-negative opportunistic pathogen in environmental and hospital settings, frequently causing respiratory diseases such as cystic fibrosis (CF), chronic obstructive pulmonary disorder (COPD), and ventilator-associated pneumonia. In our previous study, a galU-deleted clinical P. aeruginosa was found to exhibit increased susceptibility to polymyxins. The galU gene plays an important role in the biosynthesis of lipopolysaccharide (LPS) O-antigen. Here, we systematically evaluated the effects of galU deletion on the phenotype and virulence of P. aeruginosa PAO1. A galU deletion mutant was successfully constructed in P. aeruginosa PAO1 by CRISPR/Cas9, and the complementation was accomplished by pUCP18 plasmid carrying wild-type galU. The changes in phenotype, virulence, and pathogenicity were systemically studied. The results revealed that knockout of galU led to the loss of O-antigen, which affected growth, virulence, and pathogenicity through various ways in P. aeruginosa, and significantly affected the susceptibility of P. aeruginosa to polymyxins. Mechanism study suggested the involvements of quorum sensing, Entner-Doudoroff pathway, and tyrosine metabolism on bacterial virulence and antibiotic susceptibility changes after galU deletion. galU and the related pathways may serve as effective targets for the treatment of P. aeruginosa infection, providing a theoretical basis for the development of novel antibacterial drugs.},
}

*Pseudomonas aeruginosa/pathogenicity/genetics/drug effects
Virulence
Animals
Pseudomonas Infections/microbiology
Anti-Bacterial Agents/pharmacology
*Gene Deletion
*Bacterial Proteins/genetics/metabolism
Phenotype
O Antigens/genetics/biosynthesis
Polymyxins/pharmacology
Quorum Sensing
Microbial Sensitivity Tests
Mice
CRISPR-Cas Systems

@article {pmid42508244,
year = {2026},
author = {Chen, W and Cao, L and Zhu, X and Kang, W and Qiu, Y and Nie, Z and Huang, Y and Lei, C},
title = {Proteolysis-triggered CRISPR/Cas activation via cascaded enzyme switches for viral protease detection.},
journal = {Biosensors & bioelectronics},
volume = {312},
number = {},
pages = {119073},
doi = {10.1016/j.bios.2026.119073},
pmid = {42508244},
issn = {1873-4235},
abstract = {Sensitive analysis of viral proteases is crucial for early infection diagnosis and antiviral drug development; however, developing activity-based assays with high sensitivity and broad adaptability remains a significant challenge. To address this, we developed a cascade signal amplification strategy that enhances protease detection sensitivity through rational coupling of two modular, protease-responsive enzyme switches. Upon recognition of the target protease, this upstream switch triggers the activation of a downstream, protease-responsive CRISPR/Cas12a effector, which converts the specific proteolytic event into an amplified fluorescence signal. This cascaded enzymatic amplification generates a robust signal output, achieving an order-of-magnitude improvement in detection sensitivity compared to single-stage CRISPR/Cas12a assays. The modular nature of the enzyme switches renders the system highly expandable, allowing for the specific detection of diverse viral proteases. We demonstrated the platform's utility in complex biological samples by sensitively monitoring 3C protease activity within enterovirus 71-infected cells. Furthermore, dose-dependent inhibition analysis using the HRV 3C protease inhibitor rupintrivir validated the system's potential for evaluating antiviral drug efficacy. Collectively, this work establishes a versatile and modular analytical platform for the sensitive detection and functional study of viral proteases.},
}

@article {pmid42508886,
year = {2026},
author = {Tian, S and Tang, X and Zhang, S and Yao, Y and Bi, R and Li, H and Sun, X and Chen, Z},
title = {Construction of the CRISPR/Cas12a-digital integrated immunoassay technology for CD44 detection.},
journal = {Analytica chimica acta},
volume = {1417},
number = {},
pages = {346010},
doi = {10.1016/j.aca.2026.346010},
pmid = {42508886},
issn = {1873-4324},
mesh = {Humans ; Immunoassay/methods ; *Hyaluronan Receptors/blood/analysis ; *CRISPR-Cas Systems/genetics ; Limit of Detection ; },
abstract = {Digital immunoassay allows for the detection of proteins at fg/mL levels by leveraging the principles of Poisson distribution. Nevertheless, the inherent Poisson noise restricts digital sensitivity to targets exceeding 100 molecules, rendering standard digital assays incapable of detecting targets below this threshold. To overcome this constraint, we introduce an integrated CRISPR/Cas12a-digital immunoassay technology. In this system, the CRISPR/Cas12a machinery first acts as a molecular amplifier, converting each target protein into multiple enzymatic reporters. These enzyme molecules are then individually quantified via a digitized readout system, enabling ultrasensitive protein detection. By harnessing the catalytic amplification of CRISPR/Cas12a, the method achieves detection of proteins at copy numbers below 100, effectively surpassing the conventional sensitivity barrier of digital immunoassays. We validated this approach through highly selective and accurate detection of CD44 protein. A linear response was observed across a concentration range of 0.05 to 5 fg/mL, conforming to the calibration model: P(X > 0) = 0.1191c + 0.0036. The limit of detection was determined to be 0.018 fg/mL, equivalent to approximately 36 molecules of CD44. The method was further applied to quantify CD44 in plasma samples from colorectal cancer patients, demonstrating its strong potential for clinical use in early cancer diagnosis and treatment monitoring.},
}

Humans
Immunoassay/methods
*Hyaluronan Receptors/blood/analysis
*CRISPR-Cas Systems/genetics
Limit of Detection

@article {pmid42510584,
year = {2026},
author = {Yang, JW and Kim, HS and Kim, YH},
title = {Redox Regulation of Plant-Root-Knot Nematode Interactions: From ROS-Mediated Immunity to Sustainable Resistance.},
journal = {Antioxidants (Basel, Switzerland)},
volume = {15},
number = {7},
pages = {},
pmid = {42510584},
issn = {2076-3921},
abstract = {Root-knot nematodes (RKNs; Meloidogyne spp.) are among the most destructive plant parasites, causing severe yield losses in diverse crops. Reactive oxygen species (ROS), particularly superoxide radicals (O2•[-]) and hydrogen peroxide (H2O2), are central regulators of plant-RKN interactions. This review synthesizes current molecular, biochemical, genetic, transcriptomic, and translational evidence showing that the outcome of infection is determined by the spatiotemporal regulation of H2O2 rather than by ROS abundance alone. In resistant interactions, nematode perception activates PTI-associated signaling through selected cell-surface receptor complexes, including some BAK1/SERK3-associated pathways, together with BIK1, Ca[2+] signaling, and RBOHD/F, generating a sustained oxidative activity associated with salicylic acid-dependent immune signaling and reduced H2O2-scavenging capacity and coupled to hypersensitive response, lignin and callose deposition, and feeding site restriction. In susceptible interactions, RKNs deploy ROS-targeting effectors such as Mi-CRT, MjTTL5, CATLe, Mj-NEROSs, and CMII to suppress ROS production, enhance antioxidant scavenging, or weaken SA-dependent defense. Evidence from a cyst-nematode system suggests that RBOH-derived ROS can restrict excessive cell death around syncytia; whether an analogous lower-redox requirement exists in RKN-induced giant cells remains unresolved. Finally, redox-based strategies, including CRISPR/Cas editing, host-induced gene silencing, chemical priming, and biocontrol, are discussed as promising approaches for durable and sustainable nematode resistance.},
}

@article {pmid42510769,
year = {2026},
author = {Wyman, SK and Romero, Z and Heo, SJ and Navarrete, M and Krishnappa, N and Kohn, DB and Martin, DIK and Walters, MC and Boffelli, D},
title = {Oligonucleotide Synthesis Errors Are a Source of Untoward Variation in HDR-Mediated Gene Editing.},
journal = {Genes},
volume = {17},
number = {7},
pages = {},
pmid = {42510769},
issn = {2073-4425},
support = {TRAN1-09292//California Institute for Regenerative Medicine/ ; CLIN1-11497//California Institute for Regenerative Medicine/ ; CLIN2-11722//California Institute for Regenerative Medicine/ ; HL151319//National Heart Lung and Blood Institute/ ; OT3HL147741//National Heart Lung and Blood Institute/ ; },
mesh = {Humans ; *Oligonucleotides/genetics ; *Recombinational DNA Repair ; High-Throughput Nucleotide Sequencing ; Anemia, Sickle Cell/genetics/therapy ; *Gene Editing ; beta-Globins/genetics ; CRISPR-Cas Systems ; Hematopoietic Stem Cells/metabolism ; },
abstract = {Background/Objectives: Single-stranded oligonucleotides (ssODNs) are used as donor templates for therapeutic gene editing by CRISPR-Cas9 cleavage and homology-directed repair (HDR). Although ssODN sequence fidelity is critical to the safety and efficacy of editing, standard quality control methods cannot resolve individual nucleotide errors. Methods: We performed deep sequencing of ssODNs from three manufacturers and amplicons from edited hematopoietic stem/progenitor cells. Results: We find that synthesis errors are present in all ssODNs tested at rates that vary more than two-fold among manufacturers, at positions that are dependent on sequence context. These synthesis errors are propagated into the genome by HDR at frequencies proportional to their abundance in the ssODN. In our sickle cell mutation correction protocol, the most prevalent SNEs are predicted to produce benign β-globin variants, while the less frequent frameshift deletions are predicted to generate β-thalassemia-like alleles. Conclusions: Current quality control standards are insufficient to detect these errors, and deep sequencing of ssODNs should be incorporated into regulatory submissions for clinical gene editing programs.},
}

Humans
*Oligonucleotides/genetics
*Recombinational DNA Repair
High-Throughput Nucleotide Sequencing
Anemia, Sickle Cell/genetics/therapy
*Gene Editing
beta-Globins/genetics
CRISPR-Cas Systems
Hematopoietic Stem Cells/metabolism

@article {pmid42510803,
year = {2026},
author = {Gonzales, PR},
title = {Advances in Functional Genomics for Human Health.},
journal = {Genes},
volume = {17},
number = {7},
pages = {},
pmid = {42510803},
issn = {2073-4425},
mesh = {Humans ; *Genomics/methods ; Genome, Human ; },
abstract = {Cytogenomics, including karyotyping, FISH, chromosomal microarrays, and optical genome mapping, has yielded significant results for clinical phenotypes in constitutional and cancer genetics, including intellectual disability, autism spectrum disorders, dysmorphic features, and hematological and solid-tissue neoplasia. However, some of these assays have yielded results of unclear significance because the abnormalities detected were often located in intergenic regions of the genome. Because these abnormalities are within the "dark matter" of the genome, their clinical significance has been a matter of speculation. However, functional genomics can explore the clinical implications of such abnormalities more robustly, whether the abnormalities disrupt topologically associating domains (TADs), delete regulatory regions, etc. Some human genetic diseases associated with these intergenic abnormalities and characterized by functional genomics include preaxial polydactyly (SHH gene), Pierre Robin syndrome (SOX9), and 5q14.3 microdeletion syndrome (MEF2C). While functional genomics is a broad research topic, this review focuses on prior and current efforts to leverage functional genomics within the intergenic regions for human health.},
}

Humans
*Genomics/methods
Genome, Human

@article {pmid41805199,
year = {2026},
author = {Marín-Sanz, M and Berlanga-Torres, JA and Guzmán-López, MH and Sánchez-León, S and Vallés, MP and Castillo, AM and Barro, F},
title = {Engineering ultra-low-gliadin wheat for celiac disease using an integrated RNAi, CRISPR, and doubled haploid strategy.},
journal = {Journal of experimental botany},
volume = {77},
number = {14},
pages = {4417-4434},
doi = {10.1093/jxb/erag131},
pmid = {41805199},
issn = {1460-2431},
support = {QUAL21_023 IAS//Junta de Andalucı́a/ ; //Conexión TRIGO, grant number 202490E049)./ ; },
mesh = {*Triticum/genetics/metabolism ; *Gliadin/genetics/metabolism ; *RNA Interference ; Haploidy ; *Celiac Disease/genetics ; *Gene Editing ; CRISPR-Cas Systems ; Glutens ; },
abstract = {The growing prevalence of gluten-related disorders in humans has driven the development of wheat varieties with reduced immunogenic gluten. This study aimed to integrate RNA interference (RNAi) and CRISPR genome editing within a doubled haploid (DH) platform to overcome challenges of gene redundancy and polyploidy in wheat gliadins. We generated DH lines from crosses between RNAi and CRISPR lines and elite wheat cultivars, enabling stable fixation of multiple genetic modifications in a single generation. Deep sequencing analysis of α-gliadin amplicons was conducted using a custom bioinformatics pipeline optimized for complex, repetitive gene families. Gluten protein profiles were evaluated using RP-HPLC and R5 monoclonal antibody. Several DH lines presented >70% reduction in immunogenic epitopes in α-gliadins, with lines outperforming both parents. Editing frequency was influenced by sgRNA efficiency and parental background. Silencing and editing combined led to nearly depleted gliadins in some lines, often with compensatory increases in other storage proteins linked to bread-making quality, such as high-molecular-weight glutenin subunits. Kernel and specific weight traits were largely maintained. This work demonstrates that combining RNAi and CRISPR in a DH platform enables efficient, heritable reduction of immunogenic gluten, providing a viable strategy for breeding wheat lines safer for individuals with gluten-related disorders.},
}

*Triticum/genetics/metabolism
*Gliadin/genetics/metabolism
*RNA Interference
Haploidy
*Celiac Disease/genetics
*Gene Editing
CRISPR-Cas Systems
Glutens

@article {pmid41928454,
year = {2026},
author = {Tietz, SM and Brenner, K and Moyo, T and Young, PR and Vivier, MA},
title = {Optimized protocol for efficient generation, confirmation, transformation, and CRISPR editing of grapevine hairy roots.},
journal = {Journal of experimental botany},
volume = {77},
number = {14},
pages = {4352-4374},
doi = {10.1093/jxb/erag165},
pmid = {41928454},
issn = {1460-2431},
support = {//South Africa Wine/ ; UID120460//National Research Foundation (NRF)/ ; },
mesh = {*Vitis/genetics/growth & development ; *Plant Roots/genetics/growth & development ; *Transformation, Genetic ; *Gene Editing/methods ; *CRISPR-Cas Systems ; Plants, Genetically Modified/genetics ; },
abstract = {Hairy root cultures (HRCs) are powerful tools in plant biotechnology but show variable establishment efficiencies, limiting broader applications. Here, we present a standardized and optimized reference methodology for the routine generation, multiplication, and maintenance of HRCs across diverse grapevine genotypes. Our workflow evaluated three Rhizobium strains, seven grapevine cultivars (three Vitis vinifera cultivars; four Vitis rootstock hybrids), multiple explant types, infection protocols, co-cultivation times, growth media types, and anti-browning agents. The resulting protocol was effective for all grapevine genotypes and, with minor adjustments, also yielded HRCs from two other important South African plant species, namely Sutherlandia frutescens and Aspalathus linearis. Useful molecular tools were developed for transformation and selection of HRCs, including universal multiplex primers for confirmation of transformation, tested antibiotic resistance markers (kanamycin and hygromycin), and fluorescent reporters (DsRed and eyGFPuv), with DsRed found to be particularly versatile. To test the system, we overexpressed the VviMYBA1 transcription factor gene, leading to increased anthocyanin accumulation and red pigmentation in HRCs. Additionally, we achieved CRISPR/Cas9 editing of the VviPUB19 gene, the first report of CRISPR-edited grapevine HRCs. Gene editing combined with HRCs can facilitate rapid gene function studies, offering an efficient alternative or pre-screening system to whole-plant transformations, that could support advanced functional genomics and biotechnological applications in grapevine.},
}

*Vitis/genetics/growth & development
*Plant Roots/genetics/growth & development
*Transformation, Genetic
*Gene Editing/methods
*CRISPR-Cas Systems
Plants, Genetically Modified/genetics

@article {pmid42215861,
year = {2026},
author = {Liu, F and Zhang, X and Yang, Y},
title = {A comparison between CARLIN and DNA Typewriter in CRISPR-mediated lineage tracing.},
journal = {BMC bioinformatics},
volume = {27},
number = {1},
pages = {},
pmid = {42215861},
issn = {1471-2105},
support = {NCI U01-CA253553/CA/NCI NIH HHS/United States ; R01-CA251950/NH/NIH HHS/United States ; NCI U01-CA253553/CA/NCI NIH HHS/United States ; R01-CA251950/NH/NIH HHS/United States ; },
mesh = {*CRISPR-Cas Systems ; *DNA/genetics ; *Gene Editing/methods ; *Clustered Regularly Interspaced Short Palindromic Repeats ; *Cell Lineage/genetics ; },
abstract = {BACKGROUND: CARLIN and DNA Typewriter are two major breakthroughs in CRISPR-based lineage tracing technology. It is essential to understand the potential and performance of these methods in lineage tracing, which provides important guidance on experimental design.

RESULTS: In this study, we systematically compare these two strategies using a unified stochastic simulation framework with known ground-truth lineages. By explicitly modeling CRISPR editing dynamics, barcode evolution, and cell division processes, the framework enables quantitative benchmarking of lineage reconstruction accuracy across diverse experimental parameter regimes. Both methods are evaluated using multiple accuracy metrics, including Robinson-Foulds accuracy and triplet accuracy, allowing a comprehensive assessment of lineage reconstruction performance under various editing probabilities, sampling depths, and lineage lengths.

CONCLUSIONS: DNA Typewriter consistently outperforms CARLIN in lineage reconstruction accuracy when sufficient numbers of recording targets are used, particularly in more cell divisions. Sequential and ordered recording in DNA Typewriter substantially reduces ambiguity in lineage inference compared to unordered CRISPR barcode editing. CARLIN's lineage-recording potential exhausts rapidly under continuous induction, limiting its effectiveness in long-term lineage tracing. Triplet accuracy provides a more permissive and informative metric than Robinson-Foulds accuracy, especially under partial sampling scenarios.},
}

*CRISPR-Cas Systems
*DNA/genetics
*Gene Editing/methods
*Clustered Regularly Interspaced Short Palindromic Repeats
*Cell Lineage/genetics

@article {pmid42233593,
year = {2026},
author = {Zhao, W and Zheng, Z and Li, R and Xie, H and Yu, H and Zhang, Y and Wu, Y and Yang, Y and Zhang, Z and Gao, H and Li, Y and Zhang, K},
title = {In Situ Amplified Mutational mRNA Imaging Using a Spatially Confined CRISPR Nanoplatform.},
journal = {Angewandte Chemie (International ed. in English)},
volume = {65},
number = {31},
pages = {e7088080},
pmid = {42233593},
issn = {1521-3773},
support = {22377110//National Natural Science Foundation of China/ ; 82402749//National Natural Science Foundation of China/ ; U23A20531//National Natural Science Foundation of China/ ; 22122409//National Natural Science Foundation of China/ ; 252300421073//Natural Science Foundation of Henan Province/ ; 252102311025//Science and Technology Department of Henan Province/ ; 2025SGAQZ-MS-03//State Key Laboratory of Metabolic Dysregulation & Prevention and Treatment of Esophageal Cancer/ ; 261111313300//Henan Provincial Key Research and Development Program/ ; },
mesh = {*RNA, Messenger/genetics/analysis ; Humans ; Mutation ; *CRISPR-Cas Systems/genetics ; },
abstract = {Highly sensitive spatial analysis of RNA mutations is essential for understanding cellular heterogeneity and disease mechanisms. Herein, we developed an integrated CRISPR/Cas13a-based nanoprobe system for rapid detection of RNA in tissue sections (Integrated CRISPR/Cas13a-based RNA Rapid Detection, InCasRD). Unlike conventional "always-on" probes that rely on accumulated probe hybridization, InCasRD leverages the trans-cleavage activity of Cas13a to achieve spatially confined signal amplification and a high signal-to-background ratio (SBR). Using InCasRD, we achieved imaging of multiple target RNAs in tumor cells within 0.5 h of incubation, including mRNA (survivin), microRNA (miR-21), and circular RNA (circ1785). Furthermore, the engineered InCasRD system enabled mapping of RNA mutations, such as the EGFR L858R and ovarian tumor domain (OTUD) single-nucleotide variant (SNV, 23439980 G>T), in tumor tissue sections, thereby facilitating clear tumor boundary delineation. Collectively, InCasRD is a powerful, one-step tool for in situ RNA analysis with potential for diagnosis and precision medicine.},
}

*RNA, Messenger/genetics/analysis
Humans
Mutation
*CRISPR-Cas Systems/genetics

@article {pmid42335403,
year = {2026},
author = {Luo, H and Gao, J and Huang, X and Fang, Y and Huang, T and Xia, Y and Yu, Z and Cao, C and Xiong, Z},
title = {Thermo-Responsive Living Microspheroids Enable a Regenerative Living Disk-Drive System for DNA Data Storage.},
journal = {Advanced materials (Deerfield Beach, Fla.)},
volume = {38},
number = {42},
pages = {e73806},
pmid = {42335403},
issn = {1521-4095},
support = {53330200321//Tsinghua University/ ; },
mesh = {*DNA/chemistry/genetics ; *Temperature ; Plasmids/genetics ; *Information Storage and Retrieval/methods ; CRISPR-Cas Systems ; },
abstract = {DNA offers exceptional information density and long-term stability, yet its practical deployment is limited by destructive readout and the absence of a reusable, physically addressable architecture that connects nanoscale molecular information with macroscale device-level data organization. Here, we present a regenerative Living Disk-Drive system based on thermo-responsive engineered living memory microspheroids (ELMMs), in which data-encoded bacteria are encapsulated as discrete, file-level living storage units. Each ELMM contains a clonal bacterial population carrying both an information plasmid, which encodes 26 × 26 pixel icon payloads and one- to three-color intracellular fluorescent retrieval indices, and a help plasmid that enables CRISPR-Cas12a/λ-Red rewriting of the data sequence and retrieval tag. A lyophilized ELMM database forms the Living Disk, which is coupled to an Optical Retriever and desktop-scale Living Drive for closed-loop retrieval, regeneration, and database replenishment. Released bacteria regrow for downstream readout or rewriting, while a fraction is re-encapsulated into new ELMMs. The tested system retains retrieval, regrowth, and sequence recovery after four months of ambient dry storage and 13 lyophilization-rehydration cycles. Model-based performance estimates are reported only as theoretical architecture-level bounds. These results establish an experimentally bounded yet extensible architecture for physically manageable and regenerative DNA memory.},
}

*DNA/chemistry/genetics
*Temperature
Plasmids/genetics
*Information Storage and Retrieval/methods
CRISPR-Cas Systems

@article {pmid42372505,
year = {2026},
author = {Wang, R and Pan, Q and Huang, Y and Dai, W and Ping, Y and Jin, Q},
title = {Versatile hollow Ca[2+]-phenolic nanoparticles for intracellular delivery of diverse bioactive molecules and CRISPR-Cas9 genome editing.},
journal = {Biomaterials},
volume = {335},
number = {},
pages = {124407},
doi = {10.1016/j.biomaterials.2026.124407},
pmid = {42372505},
issn = {1878-5905},
mesh = {Humans ; *Nanoparticles/chemistry/ultrastructure ; *Gene Editing/methods ; *CRISPR-Cas Systems/genetics ; Animals ; *Calcium/chemistry ; *Phenols/chemistry ; },
abstract = {The development of versatile nanoplatforms capable of universally encapsulating diverse bioactive molecules holds significant promise in biomedicine. In this study, size-tunable hollow Ca[2+]-tannic acid (TA) nanoparticles (HCT NPs) are synthesized as universally applicable drug nanocarriers by simply adding TA into amorphous calcium carbonate nanoparticles. The formation of HCT NPs is identified as a surface-protected self-etching process. A wide range of hydrophobic and hydrophilic small-molecule drugs, metal ions, and biomacromolecules including proteins and nucleic acids can be encapsulated in HCT NPs for efficient intracellular delivery. HCT NPs show rapid and efficient endosomal escape, which is crucial for maintaining the bioactivity of biomacromolecules. Remarkably, a wide array of cargo proteins, spanning different molecular weights and isoelectric points can be delivered into the cytosol by HCT NPs without compromising their bioactivities. The therapeutic potential of HCT NPs for intracellular cargo delivery is exemplified by cytosolic delivery of Cas9 plasmids and Cas9 ribonucleoprotein (RNP) for CRISPR-Cas9 genome editing both in vitro and in vivo. The facile and ultrafast synthesis, versatile cargo encapsulation capabilities, efficient cell uptake and endosomal escape, and excellent biocompatibility make HCT NPs a prominent candidate for intracellular delivery of diverse bioactive molecules, particularly in therapeutic applications such as genome editing.},
}

Humans
*Nanoparticles/chemistry/ultrastructure
*Gene Editing/methods
*CRISPR-Cas Systems/genetics
Animals
*Calcium/chemistry
*Phenols/chemistry

@article {pmid42390232,
year = {2026},
author = {Duvenage, L and Chetty, A and Thomson, DD and Ballou, ER and Govender, NP and Rappleye, CA and Hoving, JC},
title = {Tools for genetic manipulation of the endemic fungal pathogen Emergomyces africanus and application of a fluorescent reporter strain in infection models.},
journal = {mSphere},
volume = {11},
number = {7},
pages = {e0018026},
pmid = {42390232},
issn = {2379-5042},
support = {209293/Z/17/Z/WT_/Wellcome Trust/United Kingdom ; 310933/Z/24/Z/WT_/Wellcome Trust/United Kingdom ; AI148561/NH/NIH HHS/United States ; },
mesh = {Animals ; Green Fluorescent Proteins/genetics ; *Genes, Reporter ; Mice ; Plasmids/genetics ; Macrophages/microbiology ; Disease Models, Animal ; *Mycoses/microbiology ; *Ascomycota/genetics/pathogenicity ; CRISPR-Cas Systems ; },
abstract = {UNLABELLED: Emergomyces africanus is a thermally dimorphic fungal pathogen endemic to Southern Africa, which can cause fatal systemic infections in persons with advanced HIV disease. Its mechanisms of pathogenesis are not well understood. Characterization of virulence traits in this pathogen requires appropriate molecular tools for genetic manipulation. Molecular technologies developed for the transformation of Histoplasma capsulatum were adapted for use in E. africanus. Agrobacterium-mediated transformation was used to generate a reporter strain expressing green fluorescent protein (GFP). The E. africanus GFP reporter strain facilitated the study of yeast interaction with macrophages in vitro and allowed the identification of infected phagocyte cell types in the mouse lung by flow cytometry. E. africanus could also maintain episomal plasmids with telomere-like sequences to introduce expression constructs without genome modification. Using this plasmid system, RNA interference constructs were used to knock down the expression of cell wall α(1,3)-glucan by targeting the transcripts of the α-glucan synthase (AGS1). An episomal CRISPR/Cas9 system was evaluated for E. africanus, which effectively disrupted GFP in a reporter strain and enabled the generation of a URA5 uracil auxotroph. These tools and strains will facilitate future studies to elucidate the mechanisms of pathogenesis of E. africanus.

IMPORTANCE: Emergomyces africanus is an opportunistic fungal pathogen affecting persons with advanced HIV disease in South Africa. The biology and pathogenesis of E. africanus are not well understood, as the importance of the disease caused by this fungus (emergomycosis) has only been recognized in recent years, and molecular studies have been impaired by the lack of genetic technologies. In this work, we describe tools and methods for the genetic modification of this pathogen, which will accelerate future studies investigating how the fungus causes disease in the human host. These essential tools include (i) the ability to create fluorescent reporter strains, such as the green fluorescent protein E. africanus strain described here, which facilitates tracking the spread of the fungus during infection and enhances microscopy studies, (ii) methods for knocking down gene expression in E. africanus, and (iii) the permanent disruption of genes through CRISPR/Cas9 gene editing.},
}

Animals
Green Fluorescent Proteins/genetics
*Genes, Reporter
Mice
Plasmids/genetics
Macrophages/microbiology
Disease Models, Animal
*Mycoses/microbiology
*Ascomycota/genetics/pathogenicity
CRISPR-Cas Systems

@article {pmid42492264,
year = {2026},
author = {Shen, H and Nourmohammadi, S and Zhou, Y and Harata-Lee, Y and Qu, Z and Wang, W and Yool, AJ and Adelson, DL},
title = {Target pathway validation for Compound Kushen Injection active against cancer cells.},
journal = {Phytomedicine : international journal of phytotherapy and phytopharmacology},
volume = {159},
number = {},
pages = {158598},
doi = {10.1016/j.phymed.2026.158598},
pmid = {42492264},
issn = {1618-095X},
abstract = {BACKGROUND: Traditional Chinese medicines are based on complex mixtures of natural products and their multi-target mechanism of action. Therefore the discovery and validation of targets and mechanisms have always been challenging. In previous studies, using transcriptomic methods and Compound Kushen Injection (CKI) as a model drug, we identified multiple pathways and candidate target genes for validation, through which CKI exerts its pharmacological effects.

PURPOSE: This study aimed to demonstrate the involvement of multiple genetic targets in different pharmacological activities for natural products in Compound Kushen Injection.

METHODS: In this study, we selected eight key genes from four candidate pathways and used CRISPR/CAS technology to knock out these genes in four cell lines, validating their role in CKI activity.

RESULTS: Although the sensitivity of different cell lines to gene knockout varied, overall, it led to reductions in various cellular activities. After the addition of CKI, we observed that, except for the minor impact of CDKN1A gene knockout on the effect of CKI, knocking out the other genes significantly affected the pharmacological efficacy of CKI in different assays. Among them, knockout of MYD88 and NFkB genes enhanced the efficacy of CKI. At the same time, we found that the genes IL24 and CYP1B1 play a crucial role in CKI inhibition of tumour cell migration, and the CYP1A1 gene is critical for the cell cycle arrest induced by CKI.

CONCLUSIONS: These findings validate the results of our previous transcriptomic analysis and further demonstrate the complexity of pharmacological mechanisms of multi-target synergistic action of natural product mixtures.},
}

@article {pmid42492655,
year = {2026},
author = {Patel, S and Panchal, J and Patel, A and Chauhan, H and Sharma, K and Sabara, P and Vahora, S and Shrimali, M and Shekh, S and Thakor, A and Mohapatra, S and Hati, S},
title = {Unravelling the Resistome of Carbapenem-Resistant E. coli from Bovine Mastitis via Whole-Genome Sequencing.},
journal = {Veterinary journal (London, England : 1997)},
volume = {},
number = {},
pages = {106792},
doi = {10.1016/j.tvjl.2026.106792},
pmid = {42492655},
issn = {1532-2971},
abstract = {Carbapenem-resistant Escherichia coli (CREC) poses a growing threat to public health, particularly when emerging from animal reservoirs such as dairy cattle. This study aimed to characterize CREC isolates recovered from bovine mastitis cases in Gujarat, India, using a combination of phenotypic antibiotic susceptibility testing and whole-genome sequencing (WGS). Out of 130 confirmed E. coli isolates from 790 mastitic milk samples, 33 (25.38%) were resistant to imipenem. Of these, nine exhibited multidrug-resistant (MDR), extensively drug-resistant (XDR), or pan-drug-resistant (PDR) phenotypes. WGS was performed on four representative isolates (SKN144, SKN685, SKN687, SKN926), revealing genome sizes ranging from 4.7 to 5.4Mb and GC content between 50.4% and 50.8%. Annotation identified numerous resistance determinants, including carbapenemase genes (blaNDM, blaOXA-48, blaTEM, blaCMY, blaCTX-M), aminoglycoside-modifying enzymes (APH, AAC), macrolide resistance genes (mphA, ermB), and multiple efflux pump systems (AcrAB-TolC, EmrAB, MdtEF-TolC). Functional genes associated with replication, repair, stress response, and mobile genetic elements (integrases, transposases, CRISPR-Cas) were also detected, indicating high genomic adaptability. Phenotypic testing revealed alarming resistance to key antimicrobials, including ampicillin (56.15%), amikacin (55.38%), ceftazidime (53.08%), and colistin (79.23%, including intermediate strains). Subsystem analysis highlighted metabolic versatility, defence mechanisms, and virulence-associated pathways. Phylogenetic analysis indicated that all isolates clustered within the same clade, suggesting possible clonal dissemination within the bovine population. The presence of CRISPR-Cas elements, integrases, and transposases suggests ongoing horizontal gene transfer and genome plasticity. These findings underscore the alarming prevalence of CREC in dairy environments and the urgent need for enhanced AMR surveillance, prudent antibiotic stewardship, and implementation of a One Health approach to prevent zoonotic transmission. This study contributes valuable genomic insights into livestock-associated CREC and highlights their close genomic parallels with high-risk human clinical clones.},
}

@article {pmid42493436,
year = {2026},
author = {Wang, Y and Song, H and DU, Y and Xu, T},
title = {[CRISPR-Cas9 activation screening identifies candidate chemokine regulators of ter-tiary lymphoid structure formation in bladder cancer].},
journal = {Beijing da xue xue bao. Yi xue ban = Journal of Peking University. Health sciences},
volume = {58},
number = {4},
pages = {707-715},
pmid = {42493436},
issn = {1671-167X},
mesh = {Animals ; Mice ; *Urinary Bladder Neoplasms/genetics/pathology/immunology ; Humans ; *Chemokines/genetics/metabolism ; Cell Line, Tumor ; Mice, Inbred C57BL ; *CRISPR-Cas Systems/genetics ; HEK293 Cells ; Chemokine CCL20/genetics ; },
abstract = {OBJECTIVE: To identify the cytokine genes influencing the formation of tertiary lymphoid structures (TLS) through CRISPR-Cas9 library screening, and to discover potential key regulatory molecules, providing new targets for enhancing the efficacy of bladder cancer immunotherapy.

METHODS: Based on a mouse whole-genome library, 44 chemokine-related genes were identified, and an single-guide RNA (sgRNA) library targeting these genes was designed and constructed, with three sgRNAs assigned to each gene. Using a lentiviral packaging system, the library plasmids were used to transfect HEK293T cells to generate a lentiviral library, which was then used to infect the mouse bladder cancer cell line MB49. Purinomycin selection was performed to obtain the MB49-mCherry cell line stably over-expressing chemokines. The cells were inoculated into the peritoneal cavity of C57BL/6 mice to establish a bladder cancer xenograft model, and tumor growth was monitored. Three weeks later, tumor tissue was excised, genomic DNA was extracted for high-throughput sequencing, and sgRNA enrichment was analyzed to screen for differentially expressed cytokine genes. Concurrently, immunohistochemical staining was performed to detect TLS markers CD20 and CD3, and the number, distribution, and maturity of TLS were assessed. The selected candidate genes were validated individually in vivo to further confirm their impact on TLS formation.

RESULTS: We successfully constructed a cytokine gene library containing 132 sgRNAs, covering 44 chemokine genes. Following lentiviral infection, we obtained the MB49-mCherry cell line, which stably expressed the library, and isolated dead Cas9-positive monoclonal cell lines via flow cytometry to ensure the homogeneity and reproducibility of subsequent experiments. Intratumoral tumor experiments in mice revealed that the number of TLS cells in the experimental group was significantly higher than in the control group, primarily distributed at the tumor margins. High-throughput sequencing results showed that, compared with the control group, in the experimental group, Cxcl16 sgDNA was significantly enriched, while Ccl20 and Cx3lc1 sgDNA levels decreased compared with baseline (P < 0.05). Further validation of the individual roles of each factor via intraperitoneal injection revealed that the number of TLSs in tumors decreased in the group treated with the CX3CL1 chemokine, suggesting that CX3CL1 might negatively regulate TLS formation. Immunohistochemical results showed that in the CX3CL1-treated group, the aggregation of CD20-positive B cells and CD3-positive T cells in the tumor tissue was reduced, and the TLS structure was incomplete.

CONCLUSION: Through CRISPR-Cas9 library screening combined with in vivo validation, this study successfully identified CX3CL1 as a potential negative regulator of TLS formation in bladder cancer. High CX3CL1 expression was associated with a reduction in TLS numbers, suggesting that it might exert an inhibitory role in the immune microenvironment of bladder cancer. This finding provides new clues and research directions for understanding the molecular mechanisms of TLS formation in bladder cancer. However, whether CX3CL1 can serve as an immunotherapeutic target remains to be further validated through clinical specimen analysis, multidimensional mechanistic investigation, and immunotherapy response correlation studies.},
}

Animals
Mice
*Urinary Bladder Neoplasms/genetics/pathology/immunology
Humans
*Chemokines/genetics/metabolism
Cell Line, Tumor
Mice, Inbred C57BL
*CRISPR-Cas Systems/genetics
HEK293 Cells
Chemokine CCL20/genetics

@article {pmid42494101,
year = {2026},
author = {Liu, K and Ren, C and Liu, X and Qi, X and Liu, Y and Ma, S and Zhang, S and Wong, XY and Wang, X and Hu, T and Hu, C},
title = {Shared CRISPR arrays underpin type I-A/I-B coexistence.},
journal = {Protein & cell},
volume = {},
number = {},
pages = {},
doi = {10.1093/procel/pwag050},
pmid = {42494101},
issn = {1674-8018},
abstract = {CRISPR-Cas systems provide adaptive immunity in prokaryotes, yet how multiple CRISPR-Cas subtypes coexist and coordinate within a single genome remains unclear. Comparative genomic analysis revealed that nearly one-third of type I-A CRISPR-Cas3 systems are adjacent to a type I-B system, often sharing a single CRISPR array. Using Thermococcus siculi RG-20 (Tsi) as a model, we show that purified TsiCas6a and TsiCas6b independently recognize and cleave the shared pre-crRNA, producing mature crRNAs with comparable efficiency. Plasmid interference assays further demonstrated that crRNAs produced by either Cas6a or Cas6b enzyme could guide both type I-A and type I-B interference complexes. This interchangeability shows that crRNAs generated by either Cas6a or Cas6b can be loaded into, and function with, both type I-A and type I-B interference complexes. Structural modelling revealed distinct but complementary recognition strategies for Cas6a and Cas6b, and mutational analysis of their RNA-binding residues impaired pre-crRNA cleavage and abolished interference activity. Together, these results uncover a shared-array logic in which Cas6a/Cas6b-compatible processing routes a single pre-crRNA to multiple type I effectors-providing a potential mechanism for subtype co-existence, a plausible explanation for array-less (or "orphan") interference modules, and an evolutionary bet-hedging strategy that prevents Acrs from shutting down immunity wholesale.},
}

@article {pmid42494113,
year = {2026},
author = {Ohtomo, M and Takarabe, S and Namiki, T and Kawata, Y and Kaneko, R and Ozawa, M and Yamada, Y and Mori, H and Kageyama, A and Kamoshita, M and Terakawa, J and Ito, J},
title = {Generation and Characterization of a Rdh1-iCre Line to Study Uterine Glandular Biology.},
journal = {Genesis (New York, N.Y. : 2000)},
volume = {64},
number = {4},
pages = {e70068},
pmid = {42494113},
issn = {1526-968X},
support = {JP21K09512//Japan Society for the Promotion of Science/ ; JP24K01950//Japan Society for the Promotion of Science/ ; JP25KJ2187//Japan Society for the Promotion of Science/ ; JP22H04922//Japan Society for the Promotion of Science/ ; JP25K22429//Japan Society for the Promotion of Science/ ; //Azabu University/ ; },
mesh = {Animals ; Female ; *Uterus/metabolism ; Mice ; *Alcohol Oxidoreductases/genetics/metabolism ; *Integrases/genetics/metabolism ; Pregnancy ; Mice, Transgenic ; CRISPR-Cas Systems ; },
abstract = {The uterus is an essential organ for fetal development in most mammals. Uterine glands, highly conserved structures in the mammalian uterus, play critical roles in the establishment and maintenance of pregnancy and have been implicated in the pathogenesis of uterine diseases, including endometrial cancer and endometriosis. Previous studies have shown that Retinol dehydrogenase 1 (Rdh1) is specifically expressed in the glandular epithelium (GE) from the onset of gland formation through adulthood. In this study, to develop a GE-specific Cre driver line, we generated Rdh1-iCre mice by introducing an improved Cre recombinase (iCre) into the Rdh1 locus using the CRISPR/Cas9 system. To evaluate the utility of this model, Rdh1-iCre mice were crossed with ROSA26-H2B-mCherry reporter mice, and Cre-dependent reporter expression was analyzed. Robust mCherry fluorescence was observed throughout the uterine glands at 2 weeks after birth, coinciding with the active elongation and branching of the GE. These results demonstrate that the Rdh1-iCre mouse line is a valuable and highly efficient tool for investigating the physiological roles of uterine glands during development and pregnancy, as well as their contribution to the progression of GE-derived uterine diseases.},
}

Animals
Female
*Uterus/metabolism
Mice
*Alcohol Oxidoreductases/genetics/metabolism
*Integrases/genetics/metabolism
Pregnancy
Mice, Transgenic
CRISPR-Cas Systems

@article {pmid42494486,
year = {2026},
author = {Wu, W and Wang, R and Li, J and Zhang, Z and Yao, W and Zhang, N and Xu, W},
title = {Engineering climate-resilient horticultural crops: advances in transcriptional regulation, genome editing, and synthetic networks.},
journal = {Horticulture research},
volume = {13},
number = {8},
pages = {uhag119},
pmid = {42494486},
issn = {2662-6810},
abstract = {Abiotic stresses-particularly cold, drought, and salinity-pose significant threats to the productivity and sustainability of horticultural crops. Recent studies have revealed conserved and species-specific regulatory mechanisms that allow plants to adapt dynamically to these environmental constraints. This review synthesizes advances in understanding key transcription factor families-such as CBF/DREB, NAC, MYB, WRKY, and bHLH-that orchestrate stress-responsive gene networks and modulate physiological processes, including osmotic regulation, antioxidant defense, and ionic homeostasis. We also discuss the emerging roles of chromatin remodeling, DNA methylation, histone modifications, and noncoding RNAs in conferring transcriptional plasticity and stress memory. Beyond endogenous pathways, we evaluate transgenic strategies, CRISPR/Cas-based genome editing, and synthetic gene circuits for engineering abiotic stress tolerance. Particular attention is given to trade-offs between growth and defense, challenges in horticultural crop transformation, and gaps in field translation. We further examine the regulatory role of secondary metabolites-such as flavonoids and salicylic acid-as biochemical interfaces between signal transduction and adaptive responses. Finally, we propose a forward-looking roadmap integrating multi-omics, ideotype design, and precision breeding toward climate-resilient horticultural systems.},
}

@article {pmid42494498,
year = {2026},
author = {Banik, I and Coppé, JP},
title = {CRISPRing through time: How cutting-edge technology is revolutionizing life sciences and medicine.},
journal = {Molecular therapy. Nucleic acids},
volume = {37},
number = {3},
pages = {103003},
pmid = {42494498},
issn = {2162-2531},
abstract = {Given the plethora of emerging technologies, none have truly captured the minds as CRISPR. From the groundbreaking research, the ultimate battle of the prizes and patents to a number of books, the science of CRISPR continues to be significant in the biomedical field. For many decades now, the emergence of synthetic biology as an intervention to correct diseases has become the foundation of biomedical research. Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based genetic editing has become a common place for routine investigation of scientific hypotheses in pre-clinical settings. More recently, CRISPR-based diagnostic testing kits for SARS-CoV-2 have showcased a translational output. Furthermore, a technological landmark was achieved when the Food and Drug Administration (FDA) approved the first CRISPR-based gene therapy (exa-cel) to edit erythroid specific enhancer region of BCL11A in hematopoietic stem cells, introduced in patients suffering from sickle cell anemia to achieve durable remission. In this review, we provide a snapshot into the most important milestones along the journey of CRISPR from its discovery in bacteria to its usage in precision medicine. The intervention of machine learning tools has now intertwined complex biology with high-throughput scalable outputs. Given the vast amount of information on CRISPR, we try to pin down key take-home messages for scientists as well as non-scientist readers. This review article attempts to understand why and how CRISPR remains significant and seamlessly integrates in the emerging era of new technologies.},
}

@article {pmid42496566,
year = {2026},
author = {Krishna, TPA and Harikrishnan, D and Veena, M and Maharajan, T and James, M and Udhayakumar, M and Arockiam Jeyasundar, PGS and David, SJ and Dineshkumar, R and Rajan, R and Rathinapriya, P},
title = {Next-Generation Metabolic Engineering of Capsaicinoids Biosynthesis in Chilli Pepper: Bridging Genomic Insights to Biotechnological Applications.},
journal = {Biotech (Basel (Switzerland))},
volume = {15},
number = {3},
pages = {},
pmid = {42496566},
issn = {2673-6284},
abstract = {Chilli peppers (Capsicum species) have been widely used around the world because of their economic value and distinctive sensory characteristics. They contain abundant functional metabolites, especially a group of vanillylamide compounds belonging to the family of capsaicinoids, which have been exploited for medicinal, nutritional, agricultural, and cosmetic uses. The demand for capsaicinoid molecules is increasing day by day due to their high economic value and wide range of applications. Therefore, increasing bioactive metabolites, especially capsaicinoids in chilli peppers, is a major priority in the current scenario. Multi-omics approaches such as genomics, transcriptomics, proteomics, and metabolomics have substantially contributed to understanding the complex regulatory networks governing capsaicinoid biosynthesis. Key structural genes, transcription factors, and signaling pathways involved in the phenylpropanoid and branched-chain fatty acid pathways have been identified, providing valuable targets for metabolic engineering in chilli pepper. Despite these advances, the integration of genetic modification approaches for the targeted enhancement of capsaicinoid production remains limited in chilli pepper. Recent developments in biotechnology, particularly CRISPR/Cas-mediated genome-editing, enable the precise genetic modification of metabolic pathways and regulatory networks in plants. Therefore, it can contribute to the precise modification of key genes involved in the capsaicinoid biosynthesis pathway, offering potential strategies to enhance the capsaicinoid content in chilli pepper. However, CRISPR/Cas-mediated genome editing in chilli pepper is still in its early stages. There are currently no reports available on the successful enhancement of capsaicinoid content in chilli peppers through CRISPR/Cas-mediated genome editing. To date, no comprehensive review has evaluated the CRISPR-Cas-mediated genome-editing approaches for capsaicinoid metabolic engineering in chilli pepper. This review critically evaluates the recent advances in CRISPR/Cas-mediated metabolic engineering in chilli peppers, with particular emphasis on regulatory genes involved in capsaicinoid biosynthesis. Furthermore, multi-omics approaches are expected to complement these strategies by enabling the identification of key regulatory genes, the optimization of genome-editing targets, and the prediction of metabolic outcomes for enhanced capsaicinoid production. Overall, this review provides insights into improving capsaicinoid accumulation in chilli peppers through advanced genome-editing technologies.},
}

@article {pmid42497250,
year = {2026},
author = {Zhou, Z and Yang, Y and Zhou, F and Liang, K and Gong, T and Zhou, X and Li, J and Luo, J and Li, J and Yang, J},
title = {Energy-assisted CRISPR cleavage and probiotic vesicle signaling platform: Microbiome reprogramming for homeostasis.},
journal = {Science advances},
volume = {12},
number = {30},
pages = {eaef1760},
pmid = {42497250},
issn = {2375-2548},
mesh = {*Probiotics ; *Homeostasis ; *Microbiota/genetics ; *Streptococcus mutans/genetics/metabolism ; Adenosine Triphosphate/metabolism ; *CRISPR-Cas Systems ; Quorum Sensing ; *Extracellular Vesicles/metabolism ; Humans ; Signal Transduction ; *Clustered Regularly Interspaced Short Palindromic Repeats ; Animals ; },
abstract = {Pathogenic infections drive microbial dysbiosis and persistent inflammation, posing therapeutic challenges due to difficulties in precise pathogen eradication and microbiome restoration. Although CRISPR-based therapeutics enable pathogen-specific antibacterial targeting, their effectiveness in treating pathogenic infections is constrained by difficulties in navigating complex microbial ecosystems, penetrating pathogenic barriers, sustaining energy-intensive intracellular cleavage, and, critically, restoring microbial balance after pathogen clearance. Here, we engineer a probiotic vesicle-synergized CRISPR platform by encapsulating gtfB-targeting CRISPR plasmids within hybrid extracellular vesicles from probiotics and pathogenic Streptococcus mutans. The pathogen-derived vesicle component enables targeted uptake by S. mutans, facilitating intracellular cleavage of the virulence gene gtfB. Vesicle-carried endogenous adenosine triphosphate (ATP) boosts CRISPR activity, amplifying targeted DNA cleavage for potent and selective pathogen elimination. Probiotic-derived vesicle components further remodel quorum-sensing networks and immunity, restoring microbial homeostasis. This probiotic vesicle-based strategy integrates ATP-enhanced CRISPR cleavage with microbiome and immune modulation, offering a next-generation therapeutic paradigm for microbiome-associated diseases.},
}

*Probiotics
*Homeostasis
*Microbiota/genetics
*Streptococcus mutans/genetics/metabolism
Adenosine Triphosphate/metabolism
*CRISPR-Cas Systems
Quorum Sensing
*Extracellular Vesicles/metabolism
Humans
Signal Transduction
*Clustered Regularly Interspaced Short Palindromic Repeats
Animals

@article {pmid42497869,
year = {2026},
author = {Terkelsen, T and Yumiceba, V and Kim, J and Melo, US and Axelgaard, E and Febbraro, F and Gunnarsson, AV and Balachandran, S and Dahl-Jessen, M and Christensen, R and Peters, BA and Asan, and Thelle, T and Nyegaard, M and Bak, RO and Denham, M and Spielmann, M and Jensen, UB},
title = {Position effect at the SOX3 locus by an interchromosomal insertion causes hereditary spastic paraplegia.},
journal = {American journal of human genetics},
volume = {},
number = {},
pages = {},
doi = {10.1016/j.ajhg.2026.07.001},
pmid = {42497869},
issn = {1537-6605},
abstract = {Pathogenic rewiring of the three-dimensional (3D) genome architecture is increasingly being identified as the cause of genetic diseases, but recognizing the cis-regulatory effects of structural variation remains a challenge. The Xq27.1 region contains a quasi-palindrome identified as a pleiotropic hotspot for disease-causing interchromosomal insertions. In a large Danish family affected by X-linked recessive complex spastic paraplegia, we identified the segregation of a 149-kb interchromosomal insertion at Xq27.1 originating from 4q24. To understand the disease mechanism, we generated induced pluripotent stem cells (iPSCs) from affected individuals. Using CRISPR perturbation and neural differentiation experiments combined with high-throughput chromatin conformation capture (Hi-C) and transcriptomic analyses, we identify a 3D regulatory rewiring of SOX3 and transcriptional dysregulation of SOX3 targets in iPSC-derived neurons. Consistent with regulatory partitioning of the SOX3 topologically associating domain (TAD) in affected individuals, our experiments show that upstream cis-regulatory elements have a reduced ability to activate SOX3 expression and that the observed dysregulation depends on CTCF-binding sites within the insertion. This work provides mechanistic evidence that a position effect at the SOX3 locus can cause hereditary spastic paraplegia.},
}

@article {pmid42500818,
year = {2026},
author = {Li, A and Zhang, X and Li, S and Wang, Y and Xu, C and Lv, C and Zhao, M and Liu, Y and Ding, M and Cao, C},
title = {ETTAS: a modular aptamer-recruited platform for programmable translational activation.},
journal = {Nucleic acids research},
volume = {54},
number = {14},
pages = {},
pmid = {42500818},
issn = {1362-4962},
support = {2021YFA0911600//National Key Research and Development Program of China/ ; A2503021//Shenzhen Medical Research Fund/ ; A2303071//Shenzhen Medical Research Fund/ ; 82403183//National Natural Science Foundation of China/ ; 82303113//National Natural Science Foundation of China/ ; 82360603//National Natural Science Foundation of China/ ; 82560157//National Natural Science Foundation of China/ ; 82300871//National Natural Science Foundation of China/ ; 82474383//National Natural Science Foundation of China/ ; JCYJ20250604180714019//Shenzhen Science and Technology Program/ ; RCJC20221008092723011//Shenzhen Science and Technology Program/ ; JCYJ20240813140522029//Shenzhen Science and Technology Program/ ; RCBS20231211090747077//Shenzhen Science and Technology Outstanding Innovative Talent Training/ ; },
mesh = {Humans ; *Aptamers, Nucleotide/genetics/metabolism ; *Protein Biosynthesis/genetics ; Tumor Suppressor Protein p53/genetics/metabolism ; *CRISPR-Cas Systems ; PTEN Phosphohydrolase/genetics/metabolism ; RNA, Guide, CRISPR-Cas Systems/genetics ; Animals ; Cell Line, Tumor ; },
abstract = {Precise enhancement of endogenous protein synthesis offers a reversible therapeutic strategy without permanent genomic modification. However, existing Cas13-mediated translational activation systems are limited by modest potency and restricted modular expandability. Here, we developed the Enhanced Targeted Translational Activation System (ETTAS), a modular RNA-guided platform that combines dCas13a, the SINEB2 translational activation element, and an independently recruitable aptamer-mediated auxiliary module. Systematic ortholog screening identified dCas13a as the most effective scaffold for SINEB2-mediated translational activation, whereas direct tandem duplication of SINEB2 elements impaired rather than enhanced activity. To overcome this architectural limitation, we used aptamer-mediated recruitment to spatially separate target recognition from auxiliary activation. A binding-validated, non-interfering dCas13a-binding aptamer enabled construction of a dual-module system in which an aptamer-recruited SINEB2 element enhanced translation without altering target mRNA abundance or stability. Compared with the previously reported dCasRx-SINEB2 system, ETTAS produced stronger reporter activation, stronger endogenous induction of P53 and PTEN, and greater antiproliferative and pro-apoptotic effects in bladder cancer cells. Proteomic analyses showed selective target protein upregulation with limited global perturbation. In vivo, dual-AAV delivery of ETTAS activated endogenous P53 and suppressed tumor growth. ETTAS establishes a programmable framework for modular post-transcriptional upregulation of endogenous proteins.},
}

Humans
*Aptamers, Nucleotide/genetics/metabolism
*Protein Biosynthesis/genetics
Tumor Suppressor Protein p53/genetics/metabolism
*CRISPR-Cas Systems
PTEN Phosphohydrolase/genetics/metabolism
RNA, Guide, CRISPR-Cas Systems/genetics
Animals
Cell Line, Tumor

@article {pmid42500819,
year = {2026},
author = {Wang, W and Chen, Y and Li, Z and Zhang, L and Gui, K and Wu, Y and Yin, N and Han, X and Zhang, Y and Lu, R and Zhang, Z and Wang, L and Xie, G},
title = {A structural accessibility principle for LbuCas13a activation by noncontiguous DNA.},
journal = {Nucleic acids research},
volume = {54},
number = {14},
pages = {},
pmid = {42500819},
issn = {1362-4962},
support = {82372351//National Natural Science Foundation of China/ ; 82572673//National Natural Science Foundation of China/ ; CSTB2024NSCQ-MSX0521//Natural Science Foundation of Chongqing/ ; CSTB2024NSCQ-MSX1223//Natural Science Foundation of Chongqing/ ; 2024M763898//China Postdoctoral Science Foundation/ ; 2024CQBSHTB3005//Chongqing Postdoctoral Special Funding Project/ ; },
mesh = {*DNA/chemistry/metabolism ; *CRISPR-Associated Proteins/metabolism/chemistry ; Models, Molecular ; Humans ; CRISPR-Cas Systems ; Molecular Dynamics Simulation ; },
abstract = {CRISPR-Cas13a is mainly known as an RNA-guided RNA endonuclease. Recent studies show that Leptotrichia buccalis Cas13a (LbuCas13a) can interact with DNA substrates too, without PAM or PFS constraints, but current understanding of DNA-mediated activation is largely based on continuous target strands. Here, we define a structural accessibility principle for LbuCas13a activation by noncontiguous DNA. We show that activation occurs only when overhang positioning creates an accessible protein-DNA interface. Outer overhangs near the crRNA repeat-adjacent side restore strong trans-cleavage activity by stabilizing key LbuCas13a-DNA contacts, whereas distal outer overhangs support only weak activation. In contrast, inner overhangs cause steric mismatch, destabilize the complex, and block formation of an active conformation. Molecular modeling and molecular dynamics simulations support this structure-dependent rule. Noncontiguous DNA also broadens the single-nucleotide discrimination window of LbuCas13a and enables accurate IDH1 R132H detection in glioma tissues. We further develop a one-step APE1-activated CRISPR-LbuCas13a reaction (ACROSS) for sensitive APE1 detection. Because activated LbuCas13a cleaves RNA reporters but not DNA-triggering products, ACROSS preserves the activating structure and supports stable signaling in vitro, in live cells, and in breast cancer serum samples.},
}

*DNA/chemistry/metabolism
*CRISPR-Associated Proteins/metabolism/chemistry
Models, Molecular
Humans
CRISPR-Cas Systems
Molecular Dynamics Simulation

@article {pmid42501080,
year = {2026},
author = {Gençoğlu, HS and Aydemir, E and Ayaz, F},
title = {Bacterial immune systems.},
journal = {Antonie van Leeuwenhoek},
volume = {119},
number = {8},
pages = {},
pmid = {42501080},
issn = {1572-9699},
mesh = {*Bacteria/immunology/virology/genetics ; Bacteriophages/immunology/physiology ; CRISPR-Cas Systems ; Immunity, Innate ; Adaptive Immunity ; },
abstract = {Bacterial immune systems encompass the multi-layered defense mechanisms that bacteria develop against bacteriophages and mobile genetic elements, such as plasmids. This review covers bacterial innate defense systems (surface defenses, superinfection exclusion, restriction-modification, abortive infection, and toxin-antitoxin systems), CRISPR-Cas-mediated adaptive immunity, and the escape strategies used by phages to overcome these defenses (genome modifications, anti-restriction proteins, and anti-CRISPR factors). Emerging evidence also highlights the role of outer membrane vesicles (OMVs) in anti-phage defense and their translational potential as vaccine and delivery platforms. In this context, a better understanding of bacterial defense systems contributes to the development of biotechnology and medical applications such as CRISPR technologies, diagnostic approaches, and phage therapy.},
}

*Bacteria/immunology/virology/genetics
Bacteriophages/immunology/physiology
CRISPR-Cas Systems
Immunity, Innate
Adaptive Immunity

@article {pmid42503339,
year = {2026},
author = {Yuan, YL and Dai, JL and Xiao, L and Jiang, JG},
title = {Advances in engineering microalgae for heterologous terpenoid synthesis: A review.},
journal = {Biotechnology advances},
volume = {},
number = {},
pages = {108993},
doi = {10.1016/j.biotechadv.2026.108993},
pmid = {42503339},
issn = {1873-1899},
abstract = {Terpenoids are a class of natural products widely distributed in living organisms, with isoprene as their fundamental structural unit. However, traditional plant extraction and chemical synthesis methods are often limited by low product purity, difficult separation, and complex synthetic steps, making it challenging to meet the demands of large-scale production. Conventional hosts such as Escherichia coli and Saccharomyces cerevisiae are utilized for terpenoid synthesis due to their advantages of short growth cycles and controllable cultivation conditions. Nevertheless, the complexity of terpenoid biosynthetic pathways poses significant challenges for these hosts in producing structurally complex terpenoids. In contrast, microalgae as photosynthetic microorganisms, possess well-developed endogenous terpenoid metabolic pathways, abundant precursor pools, and subcellular structures and regulatory mechanisms similar to those of plants, demonstrating significant advantages in the heterologous production of complex terpenoids. This review systematically summarizes recent advances in the production of heterologously synthesized terpenoids in eukaryotic microalgae, ranging from monoterpenes to triterpenes, and provides an in-depth analysis of key engineering strategies, including MEP/MVA pathway regulation, gene expression optimization, subcellular compartmentalization, and cultivation process intensification. In addition, the application potential of advanced tools such as CRISPR/Cas, microalgae-microorganism co-culture, and artificial intelligence is introduced. Finally, the major bottlenecks faced by microalgae as a sustainable green cell factory for terpenoid production are briefly analyzed, and future research directions are proposed.},
}

@article {pmid42503939,
year = {2026},
author = {Yang, XP and Gao, YQ and Huang, L and Zhao, LJ and Xiao, Y and Li, L and Zhang, TS},
title = {[Advances in research on the molecular mechanisms and gene therapy of hereditary hearing impairment].},
journal = {Zhonghua yu fang yi xue za zhi [Chinese journal of preventive medicine]},
volume = {60},
number = {7},
pages = {1138-1149},
doi = {10.3760/cma.j.cn112150-20251230-01243},
pmid = {42503939},
issn = {0253-9624},
support = {82560219//National Natural Science Foundation of China/ ; 2025S129//Kunming Medical University 2025 Master's Degree Education Innovation Fund Project/ ; },
mesh = {Humans ; *Genetic Therapy ; Connexin 26 ; *Hearing Loss/genetics/therapy ; Animals ; Connexins ; },
abstract = {Hereditary hearing impairment represents a significant etiology of language and social dysfunction in both children and adults, primarily caused by genetic factors. To date, over 150 genes have been identified in association with this disorder. The pathogenic mechanisms involve multiple molecular levels, including abnormalities in hair cell cytoskeleton and stereociliary structure, dysfunction of intercellular gap junctions (e.g., GJB2, GJB6), dysregulation of ion channels and transporters (e.g., SLC26A4, KCNQ4), alterations in extracellular matrix composition, and disruption of intracellular signaling pathways. In recent years, research has expanded to investigate the role of the inner ear immune microenvironment in this condition, with emerging evidence suggesting that immune dysregulation may contribute to disease initiation and progression. Therapeutically, novel strategies such as adeno-associated virus (AAV)-based gene replacement therapy, CRISPR/Cas-mediated gene editing systems, and lipid nanoparticle (LNP)-delivered mRNA therapeutics have demonstrated partial restoration of auditory function in animal models of hereditary hearing impairment involving genes such as TMC1, OTOF, and GJB2, with some approaches having advanced to clinical trial stages. This article systematically summarizes recent advances in the molecular mechanisms, immune microenvironment involvement, and gene therapy strategies for hereditary hearing impairment, delineates the research trajectory from gene discovery and mechanistic elucidation to therapeutic development, and discusses future translational research directions and clinical challenges.},
}

Humans
*Genetic Therapy
Connexin 26
*Hearing Loss/genetics/therapy
Animals
Connexins

@article {pmid42507199,
year = {2026},
author = {Bhadra, M and Sachan, M and Nara, S},
title = {Recent advances in nanozyme assisted miRNA biosensing for disease diagnosis.},
journal = {Mikrochimica acta},
volume = {193},
number = {8},
pages = {},
pmid = {42507199},
issn = {1436-5073},
mesh = {*MicroRNAs/analysis ; *Biosensing Techniques/methods ; Humans ; *Nanostructures/chemistry ; Electrochemical Techniques/methods ; Colorimetry/methods ; },
abstract = {MiRNAs have emerged as key biomarkers for early disease detection and therapeutic monitoring; yet, their ultrasensitive and specific detection confronts challenges due to low expression levels in biological fluids and high sequence homology, therefore it demands innovative sensing strategies. Enzyme mimicking nanostructures or nanozymes, enable manifold signal amplification and ultrasensitive analyte detection. In recent years, nanozymes, mainly oxidases and peroxidases, have been used as signal amplifiers in miRNA sensors due to their detectable catalytic products at low concentrations. Considering the increasing use of nanozymes in miRNA detection, this dedicated review presents the latest advancements in the field of miRNA diagnostics and unravels the contributions of nanozymes in miRNA sensing. This work highlights two major approaches commonly used for miRNA detection in sensors; (i) assisted through nucleic acid amplification, and (ii) amplification-free approaches. It then comprehensively underpins nanozymes as signal amplifier in both aforementioned strategies through diverse sensing modalities such as colorimetry, electrochemical, SERS, and chemiluminescent. It further presents the enhancement of sensitivity through integration of nanozymes with latest technologies like machine learning based approaches, CRISPR-Cas, or towards designing point of care sensors. Lastly, the review explores the challenges in translating nanozyme-derived miRNA sensing platforms to clinical settings.},
}

*MicroRNAs/analysis
*Biosensing Techniques/methods
Humans
*Nanostructures/chemistry
Electrochemical Techniques/methods
Colorimetry/methods

@article {pmid42507484,
year = {2026},
author = {Liang, Y and Qi, X and Gao, S and Wang, Y and Kan, G and Valentovich, LN and Guo, J and An, Y},
title = {Multi-level precise regulation of gene transcription in the yeast Saccharomyces cerevisiae based on light-sensitive CRISPR/Cas systems.},
journal = {Nucleic acids research},
volume = {54},
number = {14},
pages = {},
pmid = {42507484},
issn = {1362-4962},
support = {32571457//National Natural Science Foundation of China/ ; JYTYB2024044//Scientific Research Projects of Liaoning Provincial Department of Education/ ; LJKZ0660//Scientific Research Projects of Liaoning Provincial Department of Education/ ; //Liaoning Provincial Academic Leadership Support Program/ ; //Liaoning Provincial Enterprise Sci-Tech Specialist Assignment Program/ ; },
mesh = {*Saccharomyces cerevisiae/genetics/metabolism ; *CRISPR-Cas Systems ; *Transcription, Genetic ; *Gene Expression Regulation, Fungal ; Luminescent Proteins/genetics/metabolism ; Light ; Optogenetics/methods ; Gene Regulatory Networks ; RNA, Guide, CRISPR-Cas Systems/genetics/metabolism ; Red Fluorescent Protein ; },
abstract = {Regulation of gene transcription based on clustered regularly interspaced short palindromic repeats (CRISPR) is a powerful tool for constructing synthetic gene circuits in Saccharomyces cerevisiae. The current CRISPR-based regulatory approaches primarily focus on inhibiting the binding of dCas9 protein to single guide RNA (sgRNA) or blocking target site recognition. However, these regulation strategies are often at a single level, and their sensitivity still needs to be improved. In this study, the gene regulatory approaches at the translational and post-translational levels were integrated with optogenetic control patterns to attain very sensitive multi-level precision regulation of the dCas9 protein, thereby facilitating flexible regulation of transcription levels of target genes. This strategy was used to regulate the transcription levels of fluorescent proteins, resulting in up to 2.58-fold increase in the fluorescence intensity of mCherry compared to that without regulation. This CRISPR-based multi-level optogenetic system should be extremely helpful in understanding gene regulatory networks and in designing robust genetic circuits for synthetic biology.},
}

*Saccharomyces cerevisiae/genetics/metabolism
*CRISPR-Cas Systems
*Transcription, Genetic
*Gene Expression Regulation, Fungal
Luminescent Proteins/genetics/metabolism
Light
Optogenetics/methods
Gene Regulatory Networks
RNA, Guide, CRISPR-Cas Systems/genetics/metabolism
Red Fluorescent Protein

@article {pmid42479319,
year = {2026},
author = {Eldemir, ME and Karaca, AN and Akçelik, N and Akçelik, M},
title = {CRISPR rewired: from adaptive immunity to a global virulence control network in Salmonella.},
journal = {World journal of microbiology & biotechnology},
volume = {42},
number = {8},
pages = {},
pmid = {42479319},
issn = {1573-0972},
mesh = {Virulence/genetics ; *CRISPR-Cas Systems/genetics ; Gene Expression Regulation, Bacterial ; *Salmonella enterica/genetics/pathogenicity/immunology ; *Adaptive Immunity ; *Clustered Regularly Interspaced Short Palindromic Repeats ; Bacterial Proteins/genetics/metabolism ; Genomic Islands ; Biofilms/growth & development ; },
abstract = {The Type I-E CRISPR/Cas system in Salmonella enterica is increasingly hypothesized to function as a condition-dependent regulatory interface rather than exclusively as an adaptive immune module. Rooted in its evolutionary origins within mobile genetic elements such as casposons, this system reflects a functional transition toward influencing bacterial pathogenesis. A central hypothesis suggests that CRISPR components-specifically Cascade, Cas3, and Cas6-are integrated into core regulatory networks governing pathogenicity islands, biofilm formation, and oxidative stress adaptation. This regulatory control likely operates through a programmed deviation from the traditional immunity paradigm, where suboptimal PAM recognition or partial sequence complementarity allows Cascade to bind DNA without licensing Cas3 for lethal cleavage. Consequently, the machinery may facilitate transcriptional modulation through steric hindrance, acting as a natural CRISPR interference mechanism. Coordinated by global regulators like H-NS and LeuO in response to environmental cues such as pH fluctuations, this system effectively functions as a molecular rheostat. Collectively, these hypotheses offer a conceptual framework for novel translational strategies, including anti-CRISPR-based therapeutics and engineered evolutionary trap concepts.},
}

Virulence/genetics
*CRISPR-Cas Systems/genetics
Gene Expression Regulation, Bacterial
*Salmonella enterica/genetics/pathogenicity/immunology
*Adaptive Immunity
*Clustered Regularly Interspaced Short Palindromic Repeats
Bacterial Proteins/genetics/metabolism
Genomic Islands
Biofilms/growth & development

@article {pmid42482458,
year = {2026},
author = {Hao, J and Xie, J and Ma, K and Li, X and Chen, X and Bao, G and Hu, J and Li, G},
title = {A novel acinetobacter phage reveals altered virulence traits in phage-resistant strains.},
journal = {Virulence},
volume = {},
number = {},
pages = {2707728},
doi = {10.1080/21505594.2026.2707728},
pmid = {42482458},
issn = {2150-5608},
abstract = {Phage therapy represents a promising alternative for combating bacterial infections. This study employed an A. baumannii isolate harboring the I-F CRISPR-Cas system as a host to isolate phage and evaluate its biological characteristics. Phage-resistant mutants were screened using a double-layer agar plate assay, and the underlying molecular mechanisms were identified through whole-genome sequencing, followed by validation via gene knockout. Transcriptome sequencing was subsequently applied to alterations in the global regulatory networks of these mutants. Our results demonstrate the successful isolation of a novel myovirus, stable at 40-50 ℃, which was successfully isolated and found to utilize the capsule as its adsorption receptor. Whole-genomic analysis confirmed its taxonomic distinction from currently published phages. Investigation into the primary resistance mechanism revealed that the capsule loss, due to an insertional mutation in the UDP-glucose 4-epimerase encoding gene galE. This conclusion was further validated through targeted gene knockout of galE. This defect concurrently attenuated bacterial virulence, as demonstrated by significantly reduced lethality in the Galleria mellonella infection model and enhanced susceptibility to serum killing, while concurrently enhancing the capacity for biofilm formation. Transcriptomic profiling indicated that the ΔgalE significantly upregulated multiple biofilm-associated genes and remodeled the transcriptomic-wide regulatory. Furthermore, the combination of carbenicillin or ceftazidime with the phage exhibited a synergistic effect in vitro, effectively inhibiting biofilm formation and suppressing the emergence of phage resistance. Overall, this work characterizes a novel phage and delineates the host's biological network changes triggered by phage resistance, offering valuable insights for developing phage-based antimicrobial strategies.},
}

@article {pmid42482984,
year = {2026},
author = {Ma, C and Zhang, J and Jiang, Y and Li, X},
title = {Hybridization chain reaction-assisted CRISPR/Cas12a strategy for rapid and visual detection of Haemophilus influenzae.},
journal = {Frontiers in cellular and infection microbiology},
volume = {16},
number = {},
pages = {1844708},
pmid = {42482984},
issn = {2235-2988},
mesh = {*Haemophilus influenzae/genetics/isolation & purification ; Humans ; Sensitivity and Specificity ; *CRISPR-Cas Systems ; *Haemophilus Infections/diagnosis/microbiology ; Rapid Diagnostic Tests ; Colorimetry/methods ; *Nucleic Acid Hybridization/methods ; *Molecular Diagnostic Techniques/methods ; Benzidines ; },
abstract = {Haemophilus influenzae (H. influenzae) is a major pathogen causing community-acquired pneumonia in children, posing a serious threat to children's health. Rapid and convenient testing is needed for effective treatment. Traditional detection methods, such as bacterial culture and qPCR are cumbersome to operate, and require sophisticated instrumentation. Here, we developed a visual CRISPR/Cas detection platform that integrates the hybridization chain reaction (HCR) and horseradish peroxidase (HRP)-catalyzed 3,3',5,5'-Tetramethylbenzidine (TMB) colorimetric change, called Vi-CasHCP. The single-base recognition capability of CRISPR/Cas12a improves the specificity, while the efficiency of HCR shortens the detection time and improves the sensitivity. The peroxidase-like activity of HRP catalyzes the oxidation of TMB to oxTMB, resulting in a blue color change. Vi-CasHCP achieved a detection limit of 11.8 CFU/mL in 70 min, after Recombinase polymerase amplification. Clinical validation using 50 respiratory samples showed complete diagnostic agreement with qPCR, with 100% sensitivity, specificity, PPV, and NPV. Therefore, this platform offers rapid detection, requires no complex instruments, and has high sensitivity, providing fast and accurate diagnostic support for clinical practice, and is particularly suitable for resource-limited settings.},
}

*Haemophilus influenzae/genetics/isolation & purification
Humans
Sensitivity and Specificity
*CRISPR-Cas Systems
*Haemophilus Infections/diagnosis/microbiology
Rapid Diagnostic Tests
Colorimetry/methods
*Nucleic Acid Hybridization/methods
*Molecular Diagnostic Techniques/methods
Benzidines

@article {pmid42484920,
year = {2026},
author = {Dhall, RK and Rana, N and Kaur, G and Mandyal, SS},
title = {Genomics assisted breeding for mildew resistance in cucumber: from gene discovery to future innovations.},
journal = {Plant molecular biology},
volume = {116},
number = {4},
pages = {},
pmid = {42484920},
issn = {1573-5028},
support = {BT/Ag/CoE/PAU-GSKIG/2020-21//Ministry of Science and Technology, Department of Biotechnology, Government of India/ ; },
mesh = {*Disease Resistance/genetics ; *Cucumis sativus/genetics/microbiology ; *Plant Diseases/microbiology/genetics/immunology ; *Plant Breeding/methods ; *Genomics/methods ; Quantitative Trait Loci/genetics ; Genome, Plant ; Ascomycota ; Chromosome Mapping ; Genome-Wide Association Study ; Polymorphism, Single Nucleotide ; },
abstract = {Cucumber is an economically important vegetable crop cultivated worldwide, but its productivity is severely affected by destructive foliar diseases particularly powdery mildew and downy mildew. These pathogens cause significant yield and quality losses and the continuous emergence of new races makes disease management increasingly challenging. Conventional approaches including cultural, biological and chemical control often provide limited and short-term effectiveness. Therefore, the development of host plant resistance remains the most sustainable and environmentally sound strategy for long-term disease control. Recent advances in cucumber genomics and molecular breeding have enabled the identification of resistance-associated loci through SNP genotyping, QTL mapping, genome-wide association studies and marker-assisted selection. Furthermore, multi-omics approaches such as transcriptomics, proteomics and metabolomics combined with innovative technologies like CRISPR/Cas-mediated genome editing, genomic selection and speed breeding are transforming resistance breeding. Therefore, by integrating advanced molecular tools with omics-driven insights, this review aims to accelerate genetic gains and facilitate the development of durable, broad-spectrum mildew-resistant cucumber cultivars for sustainable and resilient production systems.},
}

*Disease Resistance/genetics
*Cucumis sativus/genetics/microbiology
*Plant Diseases/microbiology/genetics/immunology
*Plant Breeding/methods
*Genomics/methods
Quantitative Trait Loci/genetics
Genome, Plant
Ascomycota
Chromosome Mapping
Genome-Wide Association Study
Polymorphism, Single Nucleotide

@article {pmid42485704,
year = {2026},
author = {Chen, J and Zheng, H and Guan, L and Kanaherarachchi, A and Munusamy, S and Kong, J and Zhou, S and Jahani, R and Guan, X},
title = {Chemical additives-enhanced CRISPR/Cas12a-based RNA detection.},
journal = {Biosensors & bioelectronics},
volume = {312},
number = {},
pages = {119048},
doi = {10.1016/j.bios.2026.119048},
pmid = {42485704},
issn = {1873-4235},
abstract = {The CRISPR/Cas12a system has revolutionized molecular diagnostics due to its RNA-guided trans-cleavage activity, enabling programmable and highly accurate nucleic acid detection. However, most Cas12a-based assays are optimized for DNA targets, while direct RNA detection constrained by limited sensitivity, typically at the nanomolar level. Existing strategies to improve the performance of RNA analysis often rely on additional DNA activators or complex auxiliary systems. Here, we report a simple yet effective chemical additive-based strategy that overcomes these limitations. This chemical additives-enhanced CRISPR/Cas12a-based RNA detection (CARD) enables femtomolar-level RNA detection using only a single crRNA, without the need for DNA activators, reverse transcription, or strand-displacement reactions. Notably, this approach can be adapted to single-stranded DNA, enabling ssDNA detection at attomolar levels. Collectively, CARD provides a straightforward, amplification-free, and highly sensitive diagnostic framework that might be readily extended to other CRISPR/Cas systems for ultrasensitive nucleic acid diagnostics.},
}

@article {pmid42486091,
year = {2026},
author = {Regmi, M and Ma, K and Bi, C and Zhang, X and Zhao, D and Yu, L and Yang, H and Wang, N and Yang, C},
title = {Base editing for precision therapeutics.},
journal = {Cell genomics},
volume = {},
number = {},
pages = {101298},
doi = {10.1016/j.xgen.2026.101298},
pmid = {42486091},
issn = {2666-979X},
abstract = {Base editing (BE), the precise installation of single-nucleotide changes in DNA or RNA without inducing double-strand breaks, holds substantial therapeutic promise for correcting single-nucleotide variants, which constitute more than half of the known pathogenic genetic variants. Recent advances have improved base editor specificity, efficiency, and delivery, enabling clinically oriented procedures. Clinically, BE has shown early success or strong translational promise in sickle cell disease, β-thalassemia, leukemia (via CAR T and epitope engineering), hypercholesterolemia (PCSK9 and ANGPTL3), alpha-1-antitrypsin deficiency, and glycogen storage disease type Ia. Key remaining challenges include bystander editing within the activity window, residual off-target DNA and RNA editing, delivery constraints (payload size, tissue targeting, and redosing limits), immunogenicity, and the need for durable long-term safety evidence across relevant cell types and disease contexts. Continued technological refinements, careful preclinical validation, and rigorous clinical assessment will be essential to fully realize BE's transformative potential in precision medicine.},
}

@article {pmid42486099,
year = {2026},
author = {Savulescu, J and Porsdam Mann, S and Gyngell, C and Schaefer, GO},
title = {Ethics of gene therapy.},
journal = {Med (New York, N.Y.)},
volume = {},
number = {},
pages = {101225},
doi = {10.1016/j.medj.2026.101225},
pmid = {42486099},
issn = {2666-6340},
abstract = {CRISPR-Cas systems, base editing, and prime editing have made precise genetic interventions possible, and several approved therapies now treat monogenic disorders that were previously untreatable. Heritable genome editing remains ethically contested. We argue that heritable interventions should not be treated as a single category subject to uniform prohibition. We distinguish three targets: catastrophic monogenic disorders, polygenic risk reduction, and non-disease trait enhancement. For catastrophic monogenic conditions in which preimplantation selection cannot yield unaffected embryos, heritable editing is permissible, and the duty of beneficence toward future persons may require it. When the alternative is certain severe suffering or early death, the expected benefits clearly outweigh the risks. For polygenic interventions, current scientific uncertainty makes clinical application premature: predictive validity remains insufficient and pleiotropic effects are poorly understood. For enhancement, the case is weaker still. Some of its benefits are positional; the risks of social stratification are significant; and the evidence base is absent. We conclude that governance frameworks should permit what the evidence supports under stringent safeguards and prohibit what it does not. The central ethical questions concern welfare, not appeals to nature or abstract notions of dignity. Where the evidence warrants it, failing to pursue heritable gene therapy responsibly may itself be an ethical failure. We outline a translational pathway for ethical germline gene editing.},
}

@article {pmid42486981,
year = {2026},
author = {Shu, X and Wang, R and Zhou, X and Cheng, F and Ma, J and Li, Z and Li, X and Wu, T and Wu, A and Xue, Q and Liu, C and Zhao, H and Cao, X and Wang, L and Zhang, S and Zhang, Y and Li, M},
title = {CRISPR-Cas regulates expression of embedded anti-phage defence systems.},
journal = {Nature},
volume = {},
number = {},
pages = {},
pmid = {42486981},
issn = {1476-4687},
abstract = {Bacteria utilize diverse defence systems to protect against harmful foreign DNA such as bacteriophages[1,2], but how these systems coordinate with each other remains poorly understood. Here we uncover CRISIS (CRISPR-supervised immune system), a widespread regulatory paradigm whereby type I CRISPR-Cas loci embed and transcriptionally modulate diverse innate defences. Small non-canonical CRISPR RNA (crRNA)-like RNAs guide the I-C CRISPR-associated complex for antiviral defence (Cascade) effector complex to inhibit promoters of diverse immune cassettes-including composite multi-system clusters-enabling their basal expression for antiviral activity while mitigating fitness costs associated with hyperactivation, such as host growth impairment or exclusion of beneficial plasmids. When CRISPR-Cas is compromised by mutation or anti-CRISPR proteins, there is a burst in transcription of these embedded defence systems, leading to higher-level innate immunity at the expense of host fitness. Together, adaptive CRISPR-Cas systems orchestrate diverse innate immune systems into a layered defence network, comprising a prokaryotic 'immunity guard' strategy.},
}

@article {pmid42490001,
year = {2026},
author = {Feng, L and Fang, J and Xu, Y and Luo, L and Hong, B and Wang, J and Ma, Y},
title = {Functional Activity of the Lysis Protein E From Phage ID52: Dependence on SecB for Efficient Host Cell Lysis.},
journal = {Biotechnology journal},
volume = {21},
number = {7},
pages = {e70271},
pmid = {42490001},
issn = {1860-7314},
support = {XZ202601ZY0032//Science and Technology Projects of Xizang Autonomous Region, China/ ; Grant No. NERCGM-OF-20250301//Opening Foundation of National Engineering Research Center of Genetic Medicine, China/ ; 2022A1515010716//the Natural Science Foundation of Guangdong Province/ ; },
mesh = {*Bacteriophages/genetics/metabolism ; *Escherichia coli/virology/genetics/metabolism ; *Viral Proteins/metabolism/genetics/chemistry ; *Bacterial Proteins/metabolism/genetics ; *Bacteriolysis ; Molecular Docking Simulation ; Molecular Dynamics Simulation ; *Escherichia coli Proteins/metabolism/genetics ; CRISPR-Cas Systems ; },
abstract = {Single gene encoded phage lysis proteins offer a promising strategy for bacterial ghost production, yet their host-dependent regulatory mechanisms remain poorly understood. Here, we investigated the lysis protein E from phage ID52 (ID52-E), which exhibits stronger lytic activity than φX174 E. By screening ID52-E-resistant mutants, we identified a four-base insertion in secB as the genetic alteration associated with lysis resistance in BL21, and CRISPR-Cas9-mediated secB disruption confirmed that SecB is required for ID52-E-mediated lysis. Proteomic analysis revealed altered protein expression in resistant mutants despite preserved bacterial morphology. Co-immunoprecipitation and biolayer interferometry supported an apparent interaction between SecB and ID52-E, with an apparent KD of 3.541 × 10[-] [8] M under the tested 1:1 fitting model. Molecular docking, molecular dynamics simulations, mutagenesis, lysis assays, and binding measurements further implicated SecB Ala145 as a key interface residue. Together, these findings identify SecB as a host factor that facilitates ID52-E-mediated bacterial lysis and provide mechanistic insight for improving bacterial ghost production.},
}

*Bacteriophages/genetics/metabolism
*Escherichia coli/virology/genetics/metabolism
*Viral Proteins/metabolism/genetics/chemistry
*Bacterial Proteins/metabolism/genetics
*Bacteriolysis
Molecular Docking Simulation
Molecular Dynamics Simulation
*Escherichia coli Proteins/metabolism/genetics
CRISPR-Cas Systems

@article {pmid42490334,
year = {2026},
author = {Lv, Y and Kulsoom, and Jia, L and Wang, Z and Wang, F},
title = {Overcoming Liquid Biopsy Barriers: Nucleic Acid Biosensors Integrating DNA Nanotechnology and CRISPR-Cas System for Cancer Precision Theranostics.},
journal = {ACS sensors},
volume = {},
number = {},
pages = {},
doi = {10.1021/acssensors.6c01673},
pmid = {42490334},
issn = {2379-3694},
abstract = {Liquid biopsy holds immense potential for the early detection of cancer, yet its clinical utility is hindered not by the lack of available tumor-associated biomarkers but by the inadequate sensitivity and clinical robustness of current molecular diagnostic tools. Nucleic acid-based biosensors have emerged as highly programmable platforms, enabling the detection of low-abundance cancer biomarkers such as microRNAs (miRNAs), circulating tumor DNA (ctDNA), and messenger RNAs (mRNAs) in complex biological fluids. Leveraging advances in DNA nanotechnology, CRISPR-Cas-mediated RNA sensing, and chemically engineered nucleic acid analogues, these biosensors achieve attomolar-level detection through nanoscale spatial confinement and enzyme-assisted signal amplification strategies. However, their clinical translation is hindered by biological sample variability, nonspecific amplification, probe degradation, and poor reproducibility. This review analyzes the core design principles of three major biosensor categories: functional DNA nanostructures, CRISPR-Cas-based sensing systems, and synthetic analogues (PNAs, SNAs). It elucidates their structural and enzymatic optimization mechanisms, distinguishes analytical from clinical sensitivity, and addresses key liquid biopsy challenges. Finally, it outlines promising strategies for clinical translation, including microfluidic integration, artificial intelligence-assisted data analysis, and theranostic nanostructures combining diagnosis with targeted therapy. This review provides a comprehensive theoretical and technical framework for the rational design of next-generation nucleic acid biosensors and offers critical insights to bridge the gap between nanoscale engineering innovation and clinical translation, ultimately advancing the development of minimally invasive and precise cancer theranostics in precision oncology.},
}

@article {pmid42491328,
year = {2026},
author = {Hussain, A and Mojgani, N and Siddiqui, MF and Khan, MI and Ali, SA},
title = {Exploring the emerging role of CRISPR-Cas systems in probiotic development.},
journal = {Engineering microbiology},
volume = {6},
number = {3},
pages = {100279},
pmid = {42491328},
issn = {2667-3703},
abstract = {Modification of the gut microbiota by beneficial microbes can enhance an organism's lifespan, giving rise to the concept of probiotics. Probiotics are live microorganisms that provide health benefits when taken in sufficient amounts. Owing to their outstanding health benefits, probiotics have experienced rapid expansion and gained interest for the development of new applications. The exploration of microbial applications via genetic modification is currently of great interest to researchers. Genetic engineering using the clustered regularly interspaced short palindromic repeat (CRISPR)-Cas system has received considerable attention and has established applications. Owing to these enhanced properties, the CRISPR-Cas system is currently used in medicine, agriculture, food, and biotechnology. Considering the adaptive immune system in bacteria, this genetic tool is used to alter the microbial genome. Lactic acid bacteria (LAB) are widely recognized for their probiotic potential, and over 40% of LAB species contain the CRISPR-Cas system. The rising demand for probiotics and their expanding applications necessitate the enhancement of their existing characteristics. The CRISPR-Cas system, recognized for its precision, accuracy, and speed, has enabled researchers to modify the genomes of probiotics, thereby enhancing their beneficial attributes. This system can enhance probiotic properties through additive, subtractive, or modulatory mechanisms. Various approaches have been developed to improve probiotic functionalities using the CRISPR-Cas system, such as substituting slow promoters with efficient alternatives, eliminating undesirable components, boosting metabolism, and increasing tolerance levels. Furthermore, CRISPR-engineered probiotics have emerged as next-generation probiotics with enhanced properties and advanced applications across diverse fields, including the food, medicine, agriculture, and pharmaceutical sectors.},
}

@article {pmid42424384,
year = {2026},
author = {Källstig, E and Ruchti, E and Raman, M and Asadzadeh, J and Schneider, BL and McCabe, BD},
title = {Highly frequent undesired insertional mutagenesis during Drosophila genome editing.},
journal = {PLoS genetics},
volume = {22},
number = {7},
pages = {e1012192},
doi = {10.1371/journal.pgen.1012192},
pmid = {42424384},
issn = {1553-7404},
mesh = {Animals ; *Drosophila melanogaster/genetics ; *Gene Editing/methods ; *CRISPR-Cas Systems/genetics ; *Mutagenesis, Insertional/genetics ; Genome, Insect ; Recombinational DNA Repair/genetics ; Genetic Vectors/genetics ; },
abstract = {CRISPR/Cas9 based genome editing employing Homology Directed Repair (HDR) from template vector sequences is a widely used technique to enable precise insertions, deletions or modifications to genes. Here, we describe an undesired and highly frequent editing event when using conventional CRISPR/Cas9 plus HDR methods for Drosophila melanogaster germline genome editing. We find that the template vector employed for HDR repair unwantedly and commonly inserts into the genome. We observe this deviation from the desired edit at multiple genomic locations, with different HDR vectors and with multiple genome editing designs. To avoid these events, we have generated a novel HDR template vector that enables animals with these undesired insertions to be identified and excluded. Our results suggest that HDR based genome edited animals must be carefully screened for unwanted vector template genomic integration in order to avoid misleading interpretations of genome editing outcomes.},
}

Animals
*Drosophila melanogaster/genetics
*Gene Editing/methods
*CRISPR-Cas Systems/genetics
*Mutagenesis, Insertional/genetics
Genome, Insect
Recombinational DNA Repair/genetics
Genetic Vectors/genetics

@article {pmid42475178,
year = {2026},
author = {Farhangmehr, S and Braunschweig, U and Wu, M and Nabeel-Shah, S and Brown, KR and Fine, JL and Moffat, J and Blencowe, BJ},
title = {A genome-wide functional analysis of conserved intronic regions reveals essential roles for speckle-associated retained introns.},
journal = {Cell reports},
volume = {45},
number = {7},
pages = {117696},
doi = {10.1016/j.celrep.2026.117696},
pmid = {42475178},
issn = {2211-1247},
abstract = {Hundreds of human introns harbor extended regions of high evolutionary conservation that have not been previously characterized. A survey of these sequences reveals that they are associated with intron retention and enriched in genes that function in RNA processing, chromatin remodeling and neuronal biology. Using a dual CRISPR-Cas editing approach, we targeted 2,600 of these regions for deletion and observed that a subset of these perturbations affects cell growth. Many of these "fitness" sequences affect intron retention and expression levels of their host genes. Deletions in nuclear speckle-associated retained introns in the FNBP4 and DDX5 genes further cause downstream effects on cell growth-related genes and intron retention, respectively. The intronic deletion in DDX5 additionally results in the accumulation of R-loops overlapping retained introns of speckle-proximal genes. Overall, the results highlight critical and multifaceted roles of highly conserved intronic sequences in the control of gene regulation, R-loop resolution, and cell growth.},
}

@article {pmid42477308,
year = {2026},
author = {Sikandar Zaman, M and Azeem, A and Nouman, A and Khalid, A and Ghafoor, S and Zia Ul Haq, M and Aslam, MT and El-Beltagi, HS},
title = {CRISPR-Cas systems for enhancing chilling tolerance in rice: recent advances and future prospects.},
journal = {Biologia futura},
volume = {},
number = {},
pages = {},
pmid = {42477308},
issn = {2676-8607},
abstract = {Rice (Oryza sativa L.) is an important staple crop in global food security and highly vulnerable to chilling stress, which greatly affects growth, development, and yield. The conventional breeding methods for enhancing chilling tolerance face numerous problems due to the polygenic nature of chilling tolerance and genetic complexities. The present review discusses the use of CRISPR-Cas genome editing technologies as an accurate and effective approach to increasing chilling tolerance in rice. We initially describe the physiological effects of chilling stress, such as membrane fluidity impairment, inhibition of photosynthesis, nutrient imbalance, and oxidative injury, and summarize major molecular pathways and genetic materials involved in chilling tolerance. The review then outlines the recent developments in CRISPR-Cas systems, including the modes of delivery (Agrobacterium-mediated transformation, protoplast transfection, and ribonucleoprotein techniques) and how they apply to rice genome editing. The precise examination of CRISPR-based functional genomics has shown that cold-responsive genes (OsMYB30, OsWRKY76, OsAnn3, OsPRP1, and OsKASI-2) are selectively manipulated, thus contributing to a clearer understanding of their functional roles in stress signaling, membrane stability, and antioxidant defense. Moreover, we also discuss recent CRISPR strategies, including multiplex editing, transcriptional reprogramming (CRISPRa/i), and omics-guided fine-tuning of gene networks. Synthesizing latest advancements, current review establishes a conceptual framework to overcome translational challenges in CRISPR-mediated improvement of complex traits in oilseed crops, through integrating the pivotal aspects of genotype-specific delivery, multi-gene network design, field validation, and the evolving regulatory landscape. The review concludes with a reflection of gaps in research and future opportunities, with a discussion on how integrated CRISPR technologies can be used to enhance the development of climate-resistant rice varieties.},
}

@article {pmid42477873,
year = {2026},
author = {Chen, Y and Gao, XH and Vichas, A and Wang, J and Golhar, R and Neuhaus, I},
title = {ALPINE: a scalable pipeline for comprehensive classification of gene-editing outcomes from long-read amplicon sequencing.},
journal = {Bioinformatics (Oxford, England)},
volume = {42},
number = {7},
pages = {},
pmid = {42477873},
issn = {1367-4811},
mesh = {*Gene Editing/methods ; CRISPR-Cas Systems ; *High-Throughput Nucleotide Sequencing/methods ; *Software ; Humans ; *Sequence Analysis, DNA/methods ; DNA Repair ; },
abstract = {SUMMARY: CRISPR genome editing has enabled precise genetic modification for gene and cell therapies, but edits often produce heterogeneous on-target outcomes, including homology-directed repair (HDR) knock-ins, DNA repair template integrations, and structural variants. Existing tools are frequently limited to short reads or lack viral vector-specific integration categories needed for therapeutic development. Here, we present ALPINE (Amplicon Long-read Pipeline for INtegration Evaluation), a scalable and reproducible pipeline for classifying and quantifying gene-editing outcomes from long-read amplicon sequencing supporting both PacBio HiFi and Oxford Nanopore platforms. ALPINE classifies reads into 10+ categories, including DNA repair vector integration subtypes, and performs variant calling near the gene-edited site with batch, multi-sample reporting. Uniquely, ALPINE can distinguish between cells treated with multiple DNA repair vectors and identify distinct molecular features, such as inverted terminal repeats (ITRs), enabling comprehensive characterization of complex gene editing outcomes. Dual-target benchmarking on simulated datasets demonstrated high accuracy for transgene integration events. Independent validation on public crosslinked-HDR dataset confirmed ALPINE's integration detection capabilities, and application to edited T cell samples demonstrated comprehensive gene-editing outcome profiling.

AVAILABILITY: ALPINE is available under MIT license at https://github.com/Maggi-Chen/ALPINE and https://doi.org/10.5281/zenodo.20272510. All analysis scripts and visualization code used in this manuscript are available at https://github.com/Maggi-Chen/ALPINE-manuscript-analysis. Simulated datasets are deposited at Zenodo (https://doi.org/10.5281/zenodo.20260865). Public dataset PRJNA913199 is available through NCBI SRA.},
}

*Gene Editing/methods
CRISPR-Cas Systems
*High-Throughput Nucleotide Sequencing/methods
*Software
Humans
*Sequence Analysis, DNA/methods
DNA Repair

@article {pmid42478499,
year = {2026},
author = {Du, Y and Shi, Y and Chen, D and Wang, S and Su, J and Liu, K and Zhang, H and Wang, F},
title = {Gene Editing-Driven Engineering of Microbial Systems and Metabolites for Precision Medicine and Sustainable Environmental Remediation.},
journal = {Small (Weinheim an der Bergstrasse, Germany)},
volume = {},
number = {},
pages = {e74410},
doi = {10.1002/smll.74410},
pmid = {42478499},
issn = {1613-6829},
support = {BJ-2023-118//National High Level Hospital Clinical Research Funding and Fundamental Research Funds for the Central Universities/ ; T2322025//National Natural Science Foundation of China/ ; 82272161//National Natural Science Foundation of China/ ; 22125701//National Natural Science Foundation of China/ ; 52372274//National Natural Science Foundation of China/ ; 22388101//National Natural Science Foundation of China/ ; 2024YFA0919300//National Key R&D Program of China/ ; 20240101175JC//Natural Science Foundation of Jilin Province, China/ ; XF012022C0200//Xiangfu Lab Research Project/ ; },
abstract = {The engineering of microorganisms is undergoing a fundamental paradigm shift, transitioning from the construction of static cell factories to the programming of dynamically responsive living materials. However, translating molecular interventions into robust macroscopic functions requires overcoming distinct microbial-specific barriers, including delivery bottlenecks and genetic stability. In this review, we establish a unified Edit-Reprogram-Functionalize conceptual framework that systematically delineates transient genetic regulation from permanent genomic engineering. We critically examine the evolutionary trajectories of five foundational technologies: plasmid engineering, CRISPR-Cas systems, base editors, prime editors, and enzyme engineering. Rather than analyzing these toolsets in isolation, we map their convergence into an integrated engineering continuum that drives the precise synthesis of two distinct output classes: engineered living microbial materials and robust microbial metabolite-derived materials. By evaluating representative breakthroughs-from ultrasound-actuated bacterial therapeutics to ultra-tough, biosynthesized protein composites-through the strict lens of host-dependent constraints, we reveal the mechanistic principles governing successful preclinical translation. Finally, we propose an actionable roadmap centered on systemic miniaturization, closed-loop control, and multi-scale integration, providing a definitive blueprint for the next generation of precision medicine, advanced biomanufacturing, and ecological remediation.},
}

@article {pmid42248880,
year = {2026},
author = {Wang, X and Jowsey, WJ and Cheung, CY and Dickerhof, N and Chapman, CL and Taka, JRH and Hampton, MB and Bashiri, G and Gardner, PP and Fineran, PC and Cook, GM and Jackson, SA and McNeil, MB},
title = {Genome scale CRISPRi reveals both shared and strain-specific vulnerabilities in genetically diverse drug-resistant strains of Mycobacterium tuberculosis.},
journal = {Nature communications},
volume = {17},
number = {1},
pages = {},
pmid = {42248880},
issn = {2041-1723},
support = {20/459//Manatu Hauora | Health Research Council of New Zealand (HRC)/ ; 22/323//Manatu Hauora | Health Research Council of New Zealand (HRC)/ ; 22/156//Manatu Hauora | Health Research Council of New Zealand (HRC)/ ; 23/228//Manatu Hauora | Health Research Council of New Zealand (HRC)/ ; },
mesh = {*Mycobacterium tuberculosis/genetics/drug effects/metabolism/isolation & purification ; Bacterial Proteins/genetics/metabolism ; DNA-Directed RNA Polymerases/genetics ; Mutation ; *Genome, Bacterial ; Rifampin/pharmacology ; *Drug Resistance, Bacterial/genetics ; Antitubercular Agents/pharmacology ; Humans ; Genotype ; Amino Acyl-tRNA Synthetases/genetics/metabolism ; CRISPR-Cas Systems ; Microbial Sensitivity Tests ; Sulfur/metabolism ; Drug Resistance, Multiple, Bacterial/genetics ; Gene Expression Regulation, Bacterial ; },
abstract = {The global health burden caused by Mycobacterium tuberculosis is aggravated by the emergence and spread of drug resistance. Mutations that cause drug resistance can have collateral effects that increase the vulnerability of downstream pathways to inhibition. Here, using genome scale CRISPR interference we identified collateral effects associated with different drug-resistant genotypes of M. tuberculosis. We demonstrate that drug resistance generated shared vulnerabilities in several overlapping functional pathways. Most drug-resistant strains were more sensitive to tRNA synthetase knockdowns than the parental drug-sensitive strain, highlighting the potential of tRNA synthetases as high-value drug targets. Additionally, the rifampicin-resistant mutant RpoB(S450L) had increased sensitivity to the dysregulation of sulphur metabolism due to transcriptional dysregulation. This increased vulnerability did not translate to all rpoB genotypes but was linked to predicted effects on transcriptional dynamics. Amongst clinical isolates, non-synonymous mutations in sulphur metabolism genes have evolved in a geographic lineage specific manner to mitigate fitness costs associated with the collateral phenotypes of drug resistance. Combined, our findings highlight the power of functional genomics in pinpointing highly vulnerable drug targets across drug-resistant strains.},
}

*Mycobacterium tuberculosis/genetics/drug effects/metabolism/isolation & purification
Bacterial Proteins/genetics/metabolism
DNA-Directed RNA Polymerases/genetics
Mutation
*Genome, Bacterial
Rifampin/pharmacology
*Drug Resistance, Bacterial/genetics
Antitubercular Agents/pharmacology
Humans
Genotype
Amino Acyl-tRNA Synthetases/genetics/metabolism
CRISPR-Cas Systems
Microbial Sensitivity Tests
Sulfur/metabolism
Drug Resistance, Multiple, Bacterial/genetics
Gene Expression Regulation, Bacterial

@article {pmid42270637,
year = {2026},
author = {Seidel, S and Zwaans, A and Regalado, S and Choi, J and Shendure, J and Stadler, T},
title = {SciPhy: A Bayesian phylogenetic framework using sequential genetic lineage tracing data.},
journal = {Nature communications},
volume = {17},
number = {1},
pages = {},
pmid = {42270637},
issn = {2041-1723},
mesh = {Bayes Theorem ; *Phylogeny ; Animals ; *Cell Lineage/genetics ; Mice ; *Software ; Computer Simulation ; CRISPR-Cas Systems ; },
abstract = {CRISPR-based lineage tracing offers a promising avenue to decipher single-cell lineage trees, especially in organisms not amenable to microscopy. Sequential genome editing records not only genetic edits but also the order in which they occur. To leverage this enriched information, we introduce SciPhy, a simulation and inference tool implemented in BEAST 2. SciPhy utilizes a Bayesian phylogenetic approach to jointly estimate time-scaled phylogenies and cell population parameters. After validation on simulated data, we use simulated and real data from a monoclonal cell culture to benchmark SciPhy against existing methods and find that it consistently reconstructs more accurate phylogenies. Compared to UPGMA, SciPhy additionally reports uncertainty and proliferation rates. Our second example applies SciPhy to murine gastruloids, demonstrating its ability to model time-varying population dynamics in early development. Together, these results establish a phylodynamic framework for the quantitative analysis of lineage tracing data. SciPhy's codebase is publicly available at https://github.com/azwaans/SciPhy .},
}

Bayes Theorem
*Phylogeny
Animals
*Cell Lineage/genetics
Mice
*Software
Computer Simulation
CRISPR-Cas Systems

@article {pmid42473674,
year = {2026},
author = {Ananda, H and Sahana, SR and Murthy, SR and Kunnath, AN and Prashant, A and Kaifi, JT and Kiran, PK and Suvilesh, KN},
title = {Genomic innovations in cancer prevention, diagnosis, prognosis and precision therapeutics.},
journal = {Frontiers in genetics},
volume = {17},
number = {},
pages = {1828450},
pmid = {42473674},
issn = {1664-8021},
abstract = {Cancer research has undergone a transformative change with the advent of high-throughput genomic technologies. Advances in next-generation sequencing accelerated the identification of somatic and germline alterations that drive tumorigenesis enabling the transition from traditional histology-based cancer classification to molecularly informed precision oncology. Large-scale sequencing initiatives and clinical genomic profiling facilitated the development of companion diagnostic assays and targeted therapies. Beyond targeted therapies, genomic innovations have also catalyzed the emergence of novel therapeutic strategies including immunogenomics-driven immunotherapies, RNA-based therapeutics, cancer vaccines and genome editing technologies based on CRISPR-Cas systems. This review summarizes the major technological developments in cancer genomics, including sequencing platforms, transcriptomic profiling, liquid biopsy, and functional genomic screening, and highlights the utility of these innovations in discovery of actionable biomarkers and next-generation therapeutic strategies. Collectively, these advances underscore the central role of genomic technologies in driving the evolution of precision oncology toward more personalized and effective cancer treatment strategies.},
}

@article {pmid42263439,
year = {2026},
author = {Guo, B and Li, Y and Shi, M and Zhang, J and Wu, Q and Wu, X and An, R and Wang, F},
title = {A triplex-readout CRISPR-Cas12a multimodal biosensing platform for point-of-care detection of avian influenza H5N1.},
journal = {Journal of hazardous materials},
volume = {514},
number = {},
pages = {142562},
doi = {10.1016/j.jhazmat.2026.142562},
pmid = {42263439},
issn = {1873-3336},
mesh = {*Influenza A Virus, H5N1 Subtype/isolation & purification/genetics ; *Biosensing Techniques/methods ; *CRISPR-Cas Systems ; *Point-of-Care Systems ; Animals ; *Influenza in Birds/diagnosis/virology ; Rapid Diagnostic Tests ; RNA, Viral/analysis ; Humans ; },
abstract = {The highly pathogenic avian influenza A (H5N1) virus poses a significant zoonotic threat to public and animal health. Conventional detection methods often face limitations in complexity, time, and equipment requirements. In this study, we report a rapid and lightweight-equipment triplex diagnostic platform for H5N1 detection, integrating locked nucleic acid (LNA)-assisted target recognition, toehold-mediated strand displacement for signal amplification, and the high specificity of the CRISPR-Cas12a system. The developed assay achieved a detection limit of 3.7 × 10[2] copies/μL for H5N1 pseudovirus RNA within a 60-minute core detection process, and exhibited excellent specificity without cross-reactivity to other influenza subtypes or coronaviruses. Moreover, it successfully identified H5N1 in clinical swab samples, yielding consistent results across three independent readout formats: fluorescence, lateral flow strip, and a portable glucose meter. With advantages of low cost, rapid operation, and multimodal verification capability, this diagnostic system is well suited for point-of-care screening in resource-limited settings and represents a promising tool for frontline outbreak response.},
}

*Influenza A Virus, H5N1 Subtype/isolation & purification/genetics
*Biosensing Techniques/methods
*CRISPR-Cas Systems
*Point-of-Care Systems
Animals
*Influenza in Birds/diagnosis/virology
Rapid Diagnostic Tests
RNA, Viral/analysis
Humans

@article {pmid42320852,
year = {2026},
author = {Tadokoro, T and Liu, N and Olson, EN},
title = {Precision modification of heart failure signaling by CRISPR-Cas9 base editing.},
journal = {Journal of molecular and cellular cardiology},
volume = {217},
number = {},
pages = {89-93},
pmid = {42320852},
issn = {1095-8584},
mesh = {Humans ; *Heart Failure/genetics/therapy/metabolism ; *Signal Transduction/genetics ; *Gene Editing/methods ; Animals ; *CRISPR-Cas Systems/genetics ; },
abstract = {Heart failure remains a leading cause of morbidity and mortality worldwide, and current therapies largely focus on symptom management and slowing disease progression rather than correcting the underlying molecular abnormalities. Recent advances in genome editing technologies have created new opportunities to treat heart failure. Among these approaches, CRISPR-Cas9 base editing has emerged as a particularly promising strategy because it enables precise nucleotide conversions without introducing double-strand DNA breaks and demonstrates relatively high efficiency in vivo. While correction of disease-causing mutations by CRISPR-Cas9 base editing represents an important application of genome editing, an alternative strategy is to directly modulate key signaling pathways that drive cardiac dysfunction. Protein kinase C alpha (PKCα) functions as a key regulator of cardiac contractility and pathological remodeling. Precision editing of phosphorylation sites that control PKCα stability or activation may therefore represent an effective strategy to suppress maladaptive kinase signaling in cardiomyocytes. This concept of "precision signaling modification" may provide a broadly applicable therapeutic approach for heart failure. Similar strategies may also be applicable to other signaling molecules, including Ca[2+]/calmodulin-dependent protein kinase II delta (CaMKIIδ), and illustrate the broader potential of signaling-focused genome editing approaches. Despite these advances, several challenges remain for clinical translation, including efficient delivery of genome editing components to the adult heart, long-term safety, and potential immune responses. Continued advances in delivery technologies and genome editing platforms may ultimately enable durable, potentially one-time therapeutic interventions for heart failure.},
}

Humans
*Heart Failure/genetics/therapy/metabolism
*Signal Transduction/genetics
*Gene Editing/methods
Animals
*CRISPR-Cas Systems/genetics

@article {pmid42470562,
year = {2026},
author = {Zhong, S and Leng, Y and Yang, S},
title = {Targeted Gene Editing in Wheat During Haploid Production via Wide Hybridization with Transgenic Maize Expressing Cas9 and Guide RNA.},
journal = {Methods in molecular biology (Clifton, N.J.)},
volume = {3029},
number = {},
pages = {313-329},
pmid = {42470562},
issn = {1940-6029},
mesh = {*Triticum/genetics ; *Zea mays/genetics ; *Gene Editing/methods ; *Haploidy ; Plants, Genetically Modified/genetics ; *RNA, Guide, CRISPR-Cas Systems/genetics ; CRISPR-Cas Systems ; Hybridization, Genetic ; Genome, Plant ; },
abstract = {The clustered regularly interspersed short palindromic repeats (CRISPR)/Cas9 system is an efficient and versatile genome engineering tool, which has been widely used for targeted mutagenesis and gene functional characterization in various organisms. This system is simple because it only requires a Cas9 enzyme serving as a nuclease and guide RNA (gRNA) containing a 20-nt sequence matching the target gene. Delivery of a vector expressing Cas9 and gRNA or the preassembled Cas9/gRNA complex as a ribonucleoprotein (RNP) into plant cells for gene targeting are usually via the biolistic- or Agrobacterium-mediated approach. However, most wheat genotypes suffer from low efficiency of callus induction and plant regeneration from explants receiving the vector or RNP delivered by the biolistic- or Agrobacterium-mediated transformation method, limiting the application of genome editing systems in many commercially grown wheat varieties. Here, we describe a stepwise protocol for targeted gene editing in wheat via wide hybridization with transgenic maize expressing Cas9 and gRNA. A binary vector expressing Cas9 and gRNA is constructed and used for Agrobacterium-mediated transformation to generate transgenic maize plants, which are used to pollinate emasculated spikes of wheat varieties. After fertilization, the maize chromosomes enter the transient hybrid zygote and the transgene (T-DNA) on a maize chromosome expresses the Cas9 enzyme and gRNA, which forms an RNP complex to edit the target gene in wheat genome. After several cell divisions, maize chromosomes in the hybrid zygote are eliminated, resulting in formation of haploid wheat embryos with the target gene edited, which can be rescued by embryo culture technique to produce haploid plants. Doubled haploid (DH) wheat plants with homozygous gene mutations are developed by chromosome doubling through colchicine treatment of the haploid plants. The wheat × maize hybridization combined with the CRISPR/Cas9 system provides a one-step approach for generating DH lines with the target gene edited from any wheat genotypes of interest.},
}

*Triticum/genetics
*Zea mays/genetics
*Gene Editing/methods
*Haploidy
Plants, Genetically Modified/genetics
*RNA, Guide, CRISPR-Cas Systems/genetics
CRISPR-Cas Systems
Hybridization, Genetic
Genome, Plant

@article {pmid42470563,
year = {2026},
author = {Shi, C and Zhang, X and Zhao, Z and Sun, MX},
title = {In Vivo Rice Haploid-Induction System by an Egg Cell-Specific Peptidase Knockout.},
journal = {Methods in molecular biology (Clifton, N.J.)},
volume = {3029},
number = {},
pages = {331-344},
pmid = {42470563},
issn = {1940-6029},
mesh = {*Oryza/genetics ; *Haploidy ; *Gene Knockout Techniques/methods ; CRISPR-Cas Systems ; *Peptide Hydrolases/genetics ; Plants, Genetically Modified/genetics ; Plant Breeding/methods ; *Plant Proteins/genetics ; },
abstract = {Doubled haploid (DH) technology is a fast and convenient approach for crop breeding and genetic research. Currently, in vitro anther culture is the main method for rice haploid production. However, genotype dependence remains a major problem in the anther culture of most rice subspecies or cultivars for haploid induction. In this chapter, we describe a protocol for in vivo haploid induction in rice using egg cell-specific peptidase (ECS) knockout lines, including mutation of ECS using CRISPR-Cas9 system, selection of homozygous Osecs mutants, and identification of haploids in the offspring of Osecs by flow cytometry. The ECS mediated maternal in vivo haploid-induction system is a convenient, time-saving and labor-saving technique to produce rice DH lines.},
}

*Oryza/genetics
*Haploidy
*Gene Knockout Techniques/methods
CRISPR-Cas Systems
*Peptide Hydrolases/genetics
Plants, Genetically Modified/genetics
Plant Breeding/methods
*Plant Proteins/genetics

@article {pmid42472629,
year = {2026},
author = {Kimura, K and Tsukamoto, M and Shishida, K and Sugisaki, H and Katahira, J and Tanaka, M and Kuwamura, M and Kol, A and Okada, M and Iijima, M and Nakanishi, M and Sugiura, K and Hatoya, S},
title = {Red blood cell differentiation using canine-induced pluripotent stem cells.},
journal = {Stem cells translational medicine},
volume = {15},
number = {8},
pages = {},
pmid = {42472629},
issn = {2157-6580},
support = {JP18H02349//JSPS KAKENHI/ ; 22J14623//JSPS KAKENHI/ ; 22H02525//JSPS KAKENHI/ ; 23K23790//JSPS KAKENHI/ ; 26K01900//JSPS KAKENHI/ ; },
mesh = {Animals ; Dogs ; *Cell Differentiation ; *Induced Pluripotent Stem Cells/cytology/metabolism ; *Erythrocytes/cytology/metabolism ; Glycophorins/metabolism/genetics ; CRISPR-Cas Systems ; Green Fluorescent Proteins/metabolism ; },
abstract = {BACKGROUND: Red blood cell (RBC) transfusions are essential for treating various medical conditions, but global demand is difficult to meet due to a dwindling donor pool and compatibility issues. Pluripotent stem cells (PSCs) offer a promising alternative of blood dependent on volunteer donors for RBC production, and dogs serve as an excellent model for translational research due to their physiological and genetic similarities to humans.

METHODS: Canine induced pluripotent stem cells (ciPSCs) were differentiated toward hematopoietic and erythroid lineages. Differentiated cells were evaluated for hematopoietic marker expression, hemoglobinization, colony-forming capacity, enucleation, and hemoglobin gene expression. Glycophorin A (GYPA)-enhanced green fluorescent protein (EGFP) reporter ciPSC lines were generated using clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9-mediated genome editing to visualize GYPA expression during differentiation.

RESULTS: This study introduces a protocol for RBC differentiation using ciPSCs. We achieved generation of hemoglobinized RBCs, progressing through polychromatic and orthochromatic erythroblast-like stages. CiPSC-derived hematopoietic cells/RBCs were confirmed to have immature characteristics as determined by limited colony-forming capacities, low enucleation, and embryonic and fetal hemoglobin gene expression. Additionally, we created GYPA-EGFP reporter ciPSC lines using CRISPR-Cas9-mediated genome editing, enabling real-time visualization of GYPA expression. This innovation confirmed GYPA as a viable surface marker for ciPSC-derived RBCs.

CONCLUSION: Our findings mark an initial step toward establishing a canine PSC-based erythroid differentiation system, providing a foundation for future improvements and exploration of applications for canine PSC-derived RBCs.},
}

Animals
Dogs
*Cell Differentiation
*Induced Pluripotent Stem Cells/cytology/metabolism
*Erythrocytes/cytology/metabolism
Glycophorins/metabolism/genetics
CRISPR-Cas Systems
Green Fluorescent Proteins/metabolism

@article {pmid42464358,
year = {2026},
author = {Farooq, A and Rafique, A and Han, E and Park, SM and Kim, HS and Kim, MJ and Hussain, A and Sheeraz Ahmad, M and LaPointe, G},
title = {Interactions of antiphage defense systems in the ESKAPE pathogen plasmids.},
journal = {Mobile DNA},
volume = {},
number = {},
pages = {},
doi = {10.1186/s13100-026-00410-2},
pmid = {42464358},
issn = {1759-8753},
support = {ALLRP 566176-2021//Natural Sciences and Engineering Research Council of Canada/ ; },
abstract = {BACKGROUND: The global rise of multidrug resistant (MDR) ESKAPE pathogens represents a serious threat to antimicrobial therapy. While phage therapy has re-emerged as a promising alternative, its effectiveness may be compromised by bacterial defense systems, particularly those encoded on plasmids. Comprehensive surveillance of the distribution, diversity, and mobilome context of plasmid-encoded defense systems in ESKAPE pathogens remains key to the design of effective phage therapies.

RESULTS: We analyzed 7,330 dereplicated plasmids from ESKAPE pathogens to characterize the prevalence, diversity, and co-occurrence of plasmid-encoded antiphage defense systems. Conjugative plasmids, especially from Enterobacter spp. and K. pneumoniae, harbored the highest prevalence and diversity of defense systems. Defense-positive plasmids showed larger sizes, higher GC content, and frequent co-occurrence of resistance genes, especially from β-lactam, aminoglycoside, and sulfonamide classes, along with transposable elements such as IS6, IS3, and Tn3. Random forest and correlation analyses confirmed TEs and ARGs as dominant predictors of defense system occurrence. Network analysis revealed structured and partially conserved interactions among defense genes, TEs, and ARGs. RM and CBASS systems were frequently linked to beta-lactam and aminoglycoside resistance genes, as well as TEs such as IS6 and IS3. Recurrent associations such as RM-IS6, RM-IS1380, CBASS-IS3 and RM-OXA suggest shared horizontal transfer mechanisms.

CONCLUSIONS: Plasmid-encoded antiphage defense systems in ESKAPE pathogens are widespread, structured, and linked to ARGs and mobile genetic elements. These findings highlight the contribution of plasmids to the dissemination of phage-resistance traits, underscore the importance of the mobilome in shaping phage-resistance landscapes in multidrug-resistant pathogens, and support the incorporation of plasmid defense profiling into phage therapy design.},
}

@article {pmid42465601,
year = {2026},
author = {Kumar, S and Singh, RM and Gadhave, KR},
title = {CRISPR-Cas systems for plant virus management: detection, surveillance, and host resistance.},
journal = {Frontiers in plant science},
volume = {17},
number = {},
pages = {1804262},
pmid = {42465601},
issn = {1664-462X},
abstract = {The CRISPR-Cas system has transformed genome manipulation by enabling precise and programmable modification of genetic material. Initially developed as a genome-editing tool, CRISPR technologies have expanded from fundamental research to applied use across plant, animal, and microbial systems due to their simplicity, accuracy, and versatility. In agriculture, CRISPR-Cas9 has progressed from crop improvement to host-directed strategies conferring resistance against a broad range of plant viruses. Concurrently, the discovery of additional Cas effector proteins, particularly Cas12a and Cas13a, has enabled highly sensitive nucleic acid-based diagnostic platforms supporting rapid, field-deployable pathogen detection. Here, we present a focused synthesis integrating CRISPR-mediated host resistance engineering with CRISPR-based diagnostic surveillance within a unified framework for plant virus management. Unlike previous reviews that treat these domains independently, we emphasize their convergence in enabling early detection, real-time surveillance, and targeted intervention across the disease cycle. Cas12a-based systems, currently the most widely implemented, have been coupled with isothermal amplification and visual readouts for rapid virus detection, whereas Cas13a-based platforms offer direct RNA targeting with potential for simplified workflows, although they remain less developed. We examine key design considerations, performance characteristics, and limitations of these platforms, including challenges related to sensitivity, multiplexing, and field deployment. Finally, we highlight future directions, including vector-based detection, multiplex diagnostics, and integration of CRISPR technologies into scalable surveillance systems. Collectively, this review positions CRISPR-based genome editing and diagnostics as complementary components of a next-generation strategy for plant virus detection, surveillance, and management.},
}

@article {pmid42467248,
year = {2026},
author = {Zhong, H and Ma, Q and Wei, J and Yang, A and Liu, Y and Zhen, D},
title = {CRISPR/cas-based biosensors for point-of-care testing: a comprehensive review of signal readout strategies.},
journal = {Archives of microbiology},
volume = {208},
number = {10},
pages = {},
pmid = {42467248},
issn = {1432-072X},
support = {202510555097//National College Students Innovation and Entrepreneurship Training Program/ ; 82503565//National Natural Science Foundation of China/ ; },
mesh = {*Biosensing Techniques/methods/instrumentation ; *CRISPR-Cas Systems ; *Point-of-Care Testing ; Humans ; Colorimetry/methods ; Rapid Diagnostic Tests ; Point-of-Care Systems ; Electrochemical Techniques/methods ; },
abstract = {The CRISPR/Cas system has emerged as a transformative tool for nucleic acid detection, offering significant potential for point-of-care testing (POCT). However, translating CRISPR/Cas-based assays into practical POCT devices critically depends on the development of portable, sensitive, and user-friendly signal readout modalities. This review systematically compares four major readout modalities: fluorescence, electrochemical, colorimetric, and distance‑based readout, analyzing their mechanisms, analytical performance, and practical limitations. Key challenges, including sample preparation, amplification-free detection, multiplexing, and commercialization barriers, are critically assessed. Finally, future perspectives are proposed: integrating microfluidics with smartphone‑based readout, leveraging artificial intelligence and the Internet of Things for automated signal interpretation and cloud connectivity, and establishing regulatory pathways for clinical translation. This review aims to provide actionable insights for researchers developing next‑generation CRISPR diagnostics and to accelerate the transition from laboratory prototypes to deployable POCT devices.},
}

*Biosensing Techniques/methods/instrumentation
*CRISPR-Cas Systems
*Point-of-Care Testing
Humans
Colorimetry/methods
Rapid Diagnostic Tests
Point-of-Care Systems
Electrochemical Techniques/methods

@article {pmid42469036,
year = {2026},
author = {Portell-Montserrat, J and Höpfler, M},
title = {Nascent peptides emerge as regulators of mRNA stability.},
journal = {Trends in cell biology},
volume = {},
number = {},
pages = {},
doi = {10.1016/j.tcb.2026.06.010},
pmid = {42469036},
issn = {1879-3088},
abstract = {Mobile genetic elements and their hosts engage in continuous evolutionary conflict. Marino et al. recently uncovered an unusual anti-CRISPR mechanism: the phage protein AcrVA2 triggers translation-coupled mRNA degradation by recognizing nascent Cas12. The findings suggest that nascent peptides may signal an underappreciated layer of gene regulation across the kingdoms of life.},
}

@article {pmid42469498,
year = {2026},
author = {Khan, T and Abro, AA and Zulfiqar, U and Alotaibi, MS and Asadullaeva, D and Allaberdiev, R and Tang, X and Fan, G},
title = {CRISPR-enabled functional genomics for bolstering plant tolerance to abiotic and biotic stress; a comprehensive review.},
journal = {Functional & integrative genomics},
volume = {26},
number = {1},
pages = {},
pmid = {42469498},
issn = {1438-7948},
mesh = {*Stress, Physiological/genetics ; *CRISPR-Cas Systems ; Drought Resistance ; *Gene Editing/methods ; *Genomics ; *Crops, Agricultural/genetics ; Plants, Genetically Modified/genetics ; },
abstract = {Climate change intensifies abiotic stresses including salinity, drought, and extreme temperatures alongside biotic threats such as pathogens and insect pests, collectively undermining global crop productivity and food security. Salinity and drought alone affect 20-50% of irrigated soils, with projections indicating that nearly half of global farmland could become saline by mid-century. Conventional breeding and earlier genome editing tools zinc finger nucleases (ZFNs), and transcription activator-like effector nucleases (ZFNs, TALENs) are constrained by genetic diversity limitations, technical complexity, and slow trait deployment. The CRISPR-Cas9 system has emerged as a transformative platform offering superior precision, efficiency, scalability, and affordability for crop improvement. This review systematically examines how CRISPR-Cas9 enables targeted engineering of stress tolerance in major crops (rice, wheat, maize, tomato, barley) through gene knockout and knock-in strategies. Key applications include editing transcription factors (ART1, DRO1, OsDST) for drought and salinity tolerance, modifying transporter genes (OsHMA2, OsNramp5) for heavy metal detoxification, and disrupting susceptibility genes (MLO, OsERF922, CsLOB1) for broad-spectrum disease and pest resistance. Beyond direct editing, we highlight emerging synergies with functional genomics, multi-omics integration, and high-throughput phenotyping to accelerate target discovery and validation. A central focus is placed on nanobiotechnology-enabled CRISPR delivery systems, including lipid nanoparticles (LNPs), exosomes, and engineered nanocarriers that overcome the plant cell wall barrier a major bottleneck in plant genetic transformation. These platforms enable efficient, genotype-independent delivery of ribonucleoprotein (RNP) complexes, facilitating DNA-free editing for sustainable crop protection. By integrating CRISPR-based precision with advances in nanodelivery and molecular breeding, this review outlines a road-map for developing climate-resilient, high-yielding, and nutritionally enhanced crops to safeguard global agricultural sustainability.},
}

*Stress, Physiological/genetics
*CRISPR-Cas Systems
Drought Resistance
*Gene Editing/methods
*Genomics
*Crops, Agricultural/genetics
Plants, Genetically Modified/genetics

@article {pmid42470537,
year = {2026},
author = {Sisay, T and Berhan, A and Mihrete, K and Hunie, E and Bizuye, A},
title = {Genomic regulation of the diphtheria toxin gene and Its implications for molecular diagnostics and surveillance in low-resource settings.},
journal = {Molecular biology reports},
volume = {53},
number = {1},
pages = {},
pmid = {42470537},
issn = {1573-4978},
mesh = {*Diphtheria Toxin/genetics/metabolism ; *Corynebacterium diphtheriae/genetics/pathogenicity ; Humans ; *Diphtheria/diagnosis/genetics/microbiology ; Genomic Islands ; Gene Expression Regulation, Bacterial ; Bacterial Proteins/genetics/metabolism ; Genome, Bacterial ; DNA-Binding Proteins ; },
abstract = {Corynebacterium diphtheriae remains a significant, though often underestimated, public health concern, particularly in low- and middle-income countries. The pathogenicity of the disease is primarily determined by diphtheria toxin (DT), which is produced by the tox gene, a bacteriophage-associated element, and is tightly regulated by the iron-dependent transcriptional repressor DtxR, encoded by the dtxR gene. Despite extensive investigation into the molecular biology of DT, its regulation within the broader genomic organization, as well as its implications for diagnostic methods and surveillance strategies, have not yet been fully elucidated. This review consolidates existing evidence regarding the genomic context and molecular regulation of the tox gene, encompassing chromosomal organization, variability in GC content, genomic islands, and mechanisms of horizontal gene transfer. Significant attention is focused on lysogenic conversion mediated by corynephages and regulatory pathways responsive to iron. We also evaluate both established and novel molecular diagnostic approaches, including PCR, real-time PCR, sequencing technologies, and isothermal amplification methods like loop-mediated isothermal amplification (LAMP). Recent genomic discoveries, including pan-genome variation, CRISPR-Cas mechanisms, and the emergence of non-toxigenic tox-bearing strains are analyzed in relation to diagnostic precision and epidemiological surveillance. Understanding the genomic regulation and evolutionary dynamics of toxin production is essential for improving diagnostic accuracy and strengthening surveillance systems, particularly in resource-limited settings where diphtheria is often underdiagnosed and underreported.},
}

*Diphtheria Toxin/genetics/metabolism
*Corynebacterium diphtheriae/genetics/pathogenicity
Humans
*Diphtheria/diagnosis/genetics/microbiology
Genomic Islands
Gene Expression Regulation, Bacterial
Bacterial Proteins/genetics/metabolism
Genome, Bacterial
DNA-Binding Proteins

@article {pmid41570009,
year = {2026},
author = {Tay, YL and Thomson, SB and Hnatova, S and Ng, S and Teo, SR and McCallum, R and Sim, B and Tarantini, L and Tai, FL and Bollati, V and Loh, M and Hayden, MR and Leavitt, BR and Pouladi, MA},
title = {Silencing of human HTT by targeted CRISPR/dCas9-mediated epigenetic editing.},
journal = {Journal of Huntington's disease},
volume = {15},
number = {3},
pages = {399-407},
doi = {10.1177/18796397251415368},
pmid = {41570009},
issn = {1879-6400},
mesh = {Humans ; *Huntingtin Protein/genetics ; *DNA Methylation/genetics ; *Gene Silencing ; *Huntington Disease/genetics/therapy ; DNA Methyltransferase 3A ; *Epigenesis, Genetic ; *CRISPR-Cas Systems ; Epigenome Editing ; DNA (Cytosine-5-)-Methyltransferases/genetics ; },
abstract = {BackgroundGene silencing is widely recognized as a promising therapeutic approach for dominant monogenic disorders. Current silencing strategies, many of which are transient, utilize RNA interference. Gene silencing may also be achieved through directed epigenetic editing using a CRISPR/dCas9 effector fused to DNA methyltransferase 3A (dCas9-DNMT3A). We used this system to direct DNA methylation to HTT, the causal gene underlying the autosomal dominant neurodegenerative disorder Huntington's disease, to assess the translational potential of this strategy for treating a genetic neurological disease.ObjectiveTo characterize the regulatory effect of targeted dCas9-DNMT3A-mediated DNA methylation at HTT.MethodsWe exploited DNA methylation profiles of high and low HTT-expressing tissues and targeted hypomethylated regions of HTT associated with high levels of HTT expression.ResultsDe novo DNA methylation of loci within defined upstream, promoter, intragenic and downstream regions of HTT resulted in robust, acute silencing of HTT. The best long-term silencing of HTT, which persisted up to 30 days, was observed when targeted DNA methylation was directed to the 5'UTR and promoter regions of HTT.ConclusionsHTT gene silencing may be achieved via targeted de novo DNA methylation within hypomethylated regulatory regions at the HTT locus. DNA methylation editing may be an attractive therapeutic approach for Huntington disease due to its potential for long-term silencing and reversibility.},
}

Humans
*Huntingtin Protein/genetics
*DNA Methylation/genetics
*Gene Silencing
*Huntington Disease/genetics/therapy
DNA Methyltransferase 3A
*Epigenesis, Genetic
*CRISPR-Cas Systems
Epigenome Editing
DNA (Cytosine-5-)-Methyltransferases/genetics