Enhance genome editing efficiency and specificity by a quick CRISPR/Cas9 system
Enhance genome editing efficiency and specificity by a quick CRISPR/Cas9 system
Abstract CRISPR/Cas9 is a powerful genome editing tool that has been successfully applied to a variety of species, including zebrafish. However, targeting efficiencies vary greatly at different genomic loci, the underlying causes of which were still elusive. Here we report a quick CRISPR/Cas9 system, designated as qCas9, which exhibits accelerated turnover of Cas9 protein in zebrafish. Our data showed that qCas9 significantly improved targeting efficiency, including both knock-out and knock-in in F0 embryos, and yielded higher germline transmission rate in founder screen. Importantly, qCas9 showed little to no off-target editing in zebrafish and profoundly reduced off-target effect in HEK293T cell line. In summary, our findings demonstrate that qCas9 is a simple, economic and highly effective method to improve genome editing efficiency in zebrafish embryos and also holds great potential in reducing off-target effect in mammalian cell lines.
Wang Dan、Li Jing、Gu Ying、Hu Yingying、Luo Zhou、Liu Da、Wu Jun、Shen Yue、Zhang Bo、Cheng Zhenchao、Wang Jian、Xu Xun、Xiao An、Sun Hai-Xi、Yang Huanming
BGI-Shenzhen||China National GeneBank||Guangdong Provincial Key Laboratory of Genome Read and Write||Guangdong Provincial Academician Workstation of BGI Synthetic GenomicsBGI-Shenzhen||China National GeneBank||Guangdong Provincial Key Laboratory of Genome Read and Write||Guangdong Provincial Academician Workstation of BGI Synthetic GenomicsBGI-Shenzhen||China National GeneBank||Guangdong Provincial Key Laboratory of Genome Read and Write||Guangdong Provincial Academician Workstation of BGI Synthetic GenomicsBGI-Shenzhen||China National GeneBank||Guangdong Provincial Key Laboratory of Genome Read and Write||Guangdong Provincial Academician Workstation of BGI Synthetic GenomicsBGI-Shenzhen||China National GeneBank||ICX ResearchKey Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking UniversityDepartment of Molecular Biology, University of Texas Southwestern Medical CenterBGI-Shenzhen||China National GeneBank||Guangdong Provincial Key Laboratory of Genome Read and Write||Guangdong Provincial Academician Workstation of BGI Synthetic Genomics||Shenzhen Engineering Laboratory for Innovative Molecular DiagnosticsKey Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking UniversityKey Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking UniversityBGI-Shenzhen||China National GeneBankBGI-Shenzhen||China National GeneBank||Guangdong Provincial Key Laboratory of Genome Read and Write||Guangdong Provincial Academician Workstation of BGI Synthetic GenomicsKey Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking UniversityBGI-Shenzhen||China National GeneBank||Guangdong Provincial Key Laboratory of Genome Read and Write||Guangdong Provincial Academician Workstation of BGI Synthetic GenomicsBGI-Shenzhen||China National GeneBank||Guangdong Provincial Academician Workstation of BGI Synthetic Genomics
生物科学研究方法、生物科学研究技术基础医学遗传学
Wang Dan,Li Jing,Gu Ying,Hu Yingying,Luo Zhou,Liu Da,Wu Jun,Shen Yue,Zhang Bo,Cheng Zhenchao,Wang Jian,Xu Xun,Xiao An,Sun Hai-Xi,Yang Huanming.Enhance genome editing efficiency and specificity by a quick CRISPR/Cas9 system[EB/OL].(2025-03-28)[2025-05-22].https://www.biorxiv.org/content/10.1101/708404.点此复制
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