Enhance genome editing efficiency and specificity by a quick CRISPR/Cas9 system

Hu, Y.; Luo, Z.; Li, J.; Wang, D.; Sun, H.-X.; Xiao, A.; Liu, D.; Cheng, Z.; Wu, J.; Shen, Y.; Xu, X.; Zhang, B.; Wang, J.; Gu, Y.; Yang, H.

doi:10.1101/708404

bioRxiv · 10.1101/708404

Enhance genome editing efficiency and specificity by a quick CRISPR/Cas9 system

Hu, Y.·Luo, Z.·Li, J.·Wang, D.·Sun, H.-X.·Xiao, A.·Liu, D.·Cheng, Z.·Wu, J.·Shen, Y.·Xu, X.·Zhang, B.·Wang, J.·Gu, Y.·Yang, H.

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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.

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Hu, Y., Luo, Z., Li, J., Wang, D., Sun, H.-X., Xiao, A., Liu, D., Cheng, Z., Wu, J., Shen, Y., Xu, X., Zhang, B., Wang, J., Gu, Y., Yang, H.. 2019-08-02. Enhance genome editing efficiency and specificity by a quick CRISPR/Cas9 system. https://doi.org/10.1101/708404

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