Frontiers in genome editing最新文献

{"title":"From recalcitrance to precision: a robust regeneration, transformation and targeted gene editing framework in <i>Cajanus cajan</i>.","authors":"Rachana Verma, Jyotsna Bharti, Arulprakash Thangaraj, Sonia Khan Sony, Isha Gupta, Puja Chakraborty, Rashmi Kaul, Bhupendra Rawat, R Shubhra Maithreyi, Jyoti Priya Samantaray, Sugyan Preet, Kunal Tanwar, Deepak Bhardwaj, Tanushri Kaul","doi":"10.3389/fgeed.2026.1815812","DOIUrl":"10.3389/fgeed.2026.1815812","url":null,"abstract":"<p><p>Pigeonpea (<i>Cajanus cajan</i> (L.) Millsp.; 2n = 2× = 22) is a drought-tolerant perennial grain legume commonly cultivated in India's rain-fed and dry land zones, and it is a remarkable natural source of minerals and protein worldwide. Despite decades of research, the constraints associated with tissue culture continue to hinder the genetic improvement of the crop, including the explant's inability to produce embryogenic calli, direct shoot formation, limited regeneration potential, and a lack of an effective transformation system. Moreover, traditional or molecular breeding approaches for crop improvement is time, resource, and labour-intensive. CRISPR/<i>Cas9</i>-mediated approach for targeted trait improvement has emerged as a robust technology for introducing desired genetic modifications in several crop plants. We sought to report an improved protocol for calli production, <i>in vitro</i> regeneration, and a <i>CRISPR-</i>mediated genome editing of the phytoene desaturase <i>(PDS)</i> gene via a biolistic-mediated transformation system in pigeonpea. The indigenously developed construct <i>(CcPDS_NICTK-2_pCRISPR-Cas9)</i> harboring pigeonpea codon-optimized <i>Cas9</i> and target-specific <i>sgRNA</i> was used for transformation in pigeonpea explants (embryonic axis and cotyledonary nodes). The addition of tailored growth regulators and silver nitrate to shoot-induction media boosted plant regeneration to about 86% (±0.04) and transformation efficiency to 46% (±0.04). Sequencing analysis revealed the incurred mutations in the native <i>CcPDS</i> gene, with an editing efficiency of approximately 10%. Moreover, this optimized approach can be utilized in the future to generate marker-free genome-edited plants, addressing biosafety concerns and facilitating the acceptance and commercialization of genetically improved crops.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1815812"},"PeriodicalIF":5.9,"publicationDate":"2026-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13287048/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148320868","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"CRISPR and Fanzor in sickle cell disease: current progress and future prospects.","authors":"Aisha Yousef Alhumoudi, Aminah Ghazi Alotaibi, Nada Fahad Alosaimi, Abdulrahman Alshalani, Saad M Alqahtani, Sarah M Alsaab, Basem Jahz Almutiri, Mohammed M H Albariqi","doi":"10.3389/fgeed.2026.1774014","DOIUrl":"10.3389/fgeed.2026.1774014","url":null,"abstract":"<p><p>Advancements in genome editing have established a new frontier for the treatment of various genetic diseases, including sickle cell disease (SCD). SCD, the most prevalent monogenic blood disorder, causes severe pain, organ damage, and reduced life expectancy. The recent clinical approval of clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9-based gene therapies for severe sickle cell anemia marks a significant milestone in treating genetic diseases. Despite these breakthroughs, limitations in CRISPR technology persist, requiring further innovation. Alternative approaches, such as the Fanzor (Fz) system, are being developed to complement CRISPR's capabilities. Unlike CRISPR, which is typically encoded within prokaryotic organisms, Fz is encoded in the eukaryotic genome, offering a universal RNA-guided mechanism applicable across all life kingdoms. Fz's eukaryotic origin may facilitate more efficient delivery across diverse cell types and tissues, enhancing its therapeutic potential. Here, we will review the current successes and limitations of the CRISPR technology in editing mutation associated with SCD. Additionally, we will explore the potential role of Fz as a genome-editing tool for SCD, a field where its application has not yet been studied.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1774014"},"PeriodicalIF":5.9,"publicationDate":"2026-05-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13246667/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148221159","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A high-throughput, streamlined cloning protocol to generate guide RNAs for CRISPR activation.","authors":"Shusen Zhu, Aura A Tamez González, Abdelrahman Alokda, Jeremy M Van Raamsdonk","doi":"10.3389/fgeed.2026.1787203","DOIUrl":"10.3389/fgeed.2026.1787203","url":null,"abstract":"<p><p>Caenorhabditis elegans is a powerful model for studying gene function and disease pathogenesis. While RNA interference effectively suppresses gene expression, CRISPR activation (CRISPRa) provides a robust tool for upregulating endogenous gene expression in <i>C. elegans</i>. Compared with traditional injection-based methods, feeding-based CRISPRa, similar to RNA interference, is easy, cost-effective and efficient. However, the traditional cloning workflows remain a bottleneck for high-throughput gene screening. Here, we present a pooled, one-step dual gRNA cloning protocol that enables rapid and efficient construction of gRNA expression vectors for CRISPRa. In a test case, by designing 55-mer and 54-mer primers, each containing one gRNA, we amplified and pooled 126 gRNA inserts in a single reaction pipeline. The 126 gRNA clones were completed in three pooled rounds, achieving 42%-56% coverage for each round, with remaining clones processed individually. This protocol dramatically reduces time, labor, and reagent consumption, while increasing scalability and maintaining reproducibility. It is particularly well suited for high-throughput screening of gene libraries or pathways and supports downstream applications such as phenotypic screening and lifespan analysis. This work advances CRISPRa-based functional genomics in <i>C. elegans</i> by providing a practical tool for large-scale gene activation studies.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1787203"},"PeriodicalIF":5.9,"publicationDate":"2026-05-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13230216/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148165916","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Reconstructing the complex architecture of the genome with molecular scissors: applying genome editing technology in precision medicine.","authors":"Md Nur Amin Khan, Rohit Das, Pooja Barik, Somasundaram Arumugam, Shiladitya Chattopadhyay","doi":"10.3389/fgeed.2026.1762449","DOIUrl":"10.3389/fgeed.2026.1762449","url":null,"abstract":"<p><p>Our past, present, and future are governed by the grand design of the genetic blueprint, which was mapped more than two decades back. While the eloquent design of our genome results from millions of years of evolution and has reached a near-perfect stage, unwanted flaws and mistakes in individual genomes can make their life miserable and worth intervention. The advent of technologies to manipulate the grand design and make it congenial for the individual has given new hope. Here, we discuss how gene editing technologies have progressed and how some of the technologies have become indispensable gears in precision medicine.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1762449"},"PeriodicalIF":5.9,"publicationDate":"2026-04-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13171518/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147965851","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Commentary: CRISPR-Cas9 mediated editing of starch branching enzyme, SBE2 gene in potato for enhanced resistant starch for health benefits.","authors":"Ling Yin","doi":"10.3389/fgeed.2026.1803282","DOIUrl":"10.3389/fgeed.2026.1803282","url":null,"abstract":"","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1803282"},"PeriodicalIF":5.9,"publicationDate":"2026-04-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13144004/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147846796","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Advances in gene editing for legume improvement: technologies, progress, and prospects.","authors":"Shallu Thakur, Shalini Pareek, G P Dixit, Geoffrey Meru, Alok Das","doi":"10.3389/fgeed.2026.1789952","DOIUrl":"10.3389/fgeed.2026.1789952","url":null,"abstract":"<p><p>Legumes are among the most important protein-rich crops in global agri-food systems. To meet the rising protein demand of a growing population, significant efforts are underway to enhance legume yield, nutritional quality, and resilience to environmental stresses through the manipulation of key genetic traits. Advanced technologies-including genetic engineering, gene editing, genomic selection, next-generation sequencing, single-cell genomics, and multi-omics-are accelerating legume improvement due to their high precision and efficiency. This review focuses on major gene-editing technologies, namely, CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated protein 9), TALENs (Transcription Activator-Like Effector Nucleases), ZFNs (Zinc Finger Nucleases), base editing (BE), and prime editing (PE), and their applications in key legume crops such as soybean (<i>Glycine max</i>), cowpea (<i>Vigna unguiculata</i>), chickpea (<i>Cicer arietinum</i>), groundnut (<i>Arachis hypogaea</i>), pea (<i>Pisum sativum</i>), barrel clover (<i>Medicago truncatula</i>), alfalfa (<i>Medicago sativa</i>), and <i>Lotus japonicus</i>. Among these platforms, CRISPR/Cas9 is the most widely adopted in legumes, largely due to its simplicity, versatility, and dependence on accurate genome sequence information and guide RNA (gRNA) design. Advances in next-generation sequencing and the growing availability of intuitive online gRNA design tools have streamlined CRISPR workflows, improving accessibility and precision. The present review indicates that CRISPR-P is the most used gRNA design tool in legume research, likely due to its early development for plant systems and integrated off-target prediction features. Therefore, alongside reviewing gene-editing applications, we emphasized the critical role of robust gRNA design tools as a foundation for successful genome editing. Future integration of artificial intelligence and large language models is expected to further enhance target prediction accuracy, minimize off-target effects, and enable more precise genome-editing strategies in legumes.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1789952"},"PeriodicalIF":5.9,"publicationDate":"2026-04-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13126311/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147824319","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development of <i>in planta</i> genome editing by transient expression of genome-editing tools in tomato.","authors":"Misaki Kobayashi, Na Renhu, Shu Takahashi, Seungje Choi, Haruto Watanabe, Martina Bianca Fuhrmann-Aoyagi, Hiroshi Ezura, Kenji Miura","doi":"10.3389/fgeed.2026.1777148","DOIUrl":"10.3389/fgeed.2026.1777148","url":null,"abstract":"<p><p>Two major processes are important for genome editing in plants: transformation by stable transfection, in which nucleic acids encoding genome-editing enzymes are introduced into plant cells and the regeneration of plant individuals from cells harboring mutations by genome-editing enzymes. The efficiency of transformation and regeneration by tissue culture varies across plant species, and is low in some practical crop species. <i>In planta</i> methods have been developed to exclude the need for tissue culture. However, few reports are available on methods that do not require stable transfection. Therefore, this study aimed to develop a new protocol for delivery genome editing tools that does not require transformation or tissue culture, by combining the <i>in planta</i> method with transient genome editing tools instead of stable transfection. Cas9, guide RNAs, and developmental regulators, which are factors involved in mitotic tissue induction, were transiently expressed by agroinfiltration of the stem tissue cut surfaces of tomatoes. New chimeric mutants, containing a mixture of cells with mutations introduced at or near the target sequence, were obtained. After examining conditions such as the concentration of <i>Agrobacterium</i> used for infection and post-infection treatment, we succeeded in obtaining chimeric mutants with an efficiency of 11.7%. In addition, most of the observed mutations were single base substitutions. These results indicate that the <i>in planta</i> method with transient expression of genome editing tools and induction of meristematic tissue can be used to introduce genome-edited mutations in tomatoes.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1777148"},"PeriodicalIF":5.9,"publicationDate":"2026-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13102611/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147790906","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Editorial: Genome editing strategies for augmenting crop resilience against climate change.","authors":"Wadzani Palnam Dauda, Amolkumar U Solanke, Ramesh Namdeo Pudake","doi":"10.3389/fgeed.2026.1813338","DOIUrl":"10.3389/fgeed.2026.1813338","url":null,"abstract":"","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1813338"},"PeriodicalIF":5.9,"publicationDate":"2026-03-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13039654/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147610789","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Harnessing myostatin pleiotropy for multitrait improvement via precision gene editing.","authors":"Yajun Chen, Ruiyao Yang, Yucai Yang, Qianguang Wang, Kai Yang, Man Xu","doi":"10.3389/fgeed.2026.1749445","DOIUrl":"10.3389/fgeed.2026.1749445","url":null,"abstract":"<p><p>The pursuit of sustainable livestock farming to meet the rising global protein demand has positioned myostatin (MSTN) gene editing as a key technology. However, the field's focus on the remarkable double-muscle phenotype has often overshadowed a systematic examination of its concomitant effects. The present review aims to bridge this gap by moving beyond a singular focus on productivity. First, the pleiotropic effects of MSTN gene editing on growth performance, carcass quality, and meat quality in cattle, swine, sheep, poultry, and aquatic species were comprehensively evaluated. Next, the cascading biological effects of MSTN editing on metabolic homeostasis, reproductive performance, and animal health and welfare werAAe analyzed in depth. Finally, the inherent limitations and ethical issues of current editing techniques were critically discussed, and future sustainable breeding programs aimed at balanced multitrait regulation were prospectively proposed. Ultimately, this review affirms that MSTN editing has a multiplicative effect on trait alterations; however, there is also a series of associated health challenges, which demonstrates that the technology's impact is systemic, generating a spectrum of trade-offs that are often species specific. Its responsible application therefore hinges on multitrait balancing strategies to simultaneously secure productivity and sustainability in animal agriculture.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1749445"},"PeriodicalIF":5.9,"publicationDate":"2026-03-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13018111/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147576704","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The precision strategy of human genome correction via a set of circular donor DNA and its cleaver.","authors":"Kohji Kusano, Kaoru Takizawa, Jitsutaro Kawaguchi, Isamu Hara, Toyotaka Mori","doi":"10.3389/fgeed.2026.1718252","DOIUrl":"10.3389/fgeed.2026.1718252","url":null,"abstract":"<p><p>Homologous recombination (HR) corrects a mutational sequence causing a genetic disease by replacing it with the normal sequence to restore a healthy state in humans. A targeted genomic breakage, such as that induced by CRISPR-Cas9, can trigger a copy-paste-type HR event; however, CRISPR-Cas9 more frequently induces imprecise non-homologous end-joining events, leading to one-step multiple knockout products for paralogous genes or homologous alleles, which can be considered a unique advantage. We have established a precision strategy for crossover-type HR-based gene editing, primed by intra-cellular circular donor cleavage (InCDC). The InCDC technique generates targeted duplication of the circular donor plasmid at the target locus in human cells, forming a doublet configuration comprising the donor DNA with the designed sequence and the target DNA with the original sequence, with much higher efficiency than conventional donor linearization techniques. This doublet form leads to the singlet form, resulting in retention of the designed allele. We found that the safety distance within the designed circular donor plasmid and its intra-cellular cleavage was particularly critical to protect a designed sequence from enzymatic exclusion, and we propose that InCDC technology enables precision genome editing, such as the replacement of a genetic disease-causing allele with the correctly designed allele.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":"8 ","pages":"1718252"},"PeriodicalIF":5.9,"publicationDate":"2026-03-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13013431/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147521869","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}