Use of Geminivirus for Delivery of CRISPR/Cas9 Components to Tobacco by Agro-infiltration

Kangquan Yin; Ting Han; Yule Liu

doi:10.21769/BioProtoc.2209

Use of Geminivirus for Delivery of CRISPR/Cas9 Components to Tobacco by Agro-infiltration

Bio Protoc. 2017 Apr 5;7(7):e2209. doi: 10.21769/BioProtoc.2209.

Authors

Kangquan Yin^{1

2}, Ting Han¹, Yule Liu¹

Affiliations

¹ Center for Plant Biology, MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing, China.
² State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.

Abstract

CRISPR/Cas9 system is a recently developed genome editing tool, and its power has been demonstrated in many organisms, including some plant species ( Wang et al., 2016 ). In eukaryotes, the Cas9/gRNA complexes target genome sites specifically and cleave them to produce double-strand breaks (DSBs), which can be repaired by non-homologous end joining (NHEJ) pathway ( Wang et al., 2016 ). Since NHEJ is error prone, mutations are thus generated. In plants, delivery of genome editing reagents is still challenging. In this protocol, we detail the procedure of a virus-based gRNA delivery system for CRISPR/Cas9 mediated plant genome editing (VIGE). This method offers a rapid and efficient way to deliver gRNA into plant cells, especially for those that are recalcitrant to transformation with Agrobacterium.

Keywords: CRISPR; Cas9; Geminivirus; Genome editing; Tobacco; VIGE.