pYAO-based CRISPR/Cas9 Creates High Genome Editing Efficiency in Arabidopsis

CRISPR/Cas9 system is a recently developed powerful technology for genome editing. The method has been successfully applied to genome editing in animals, and also to modify multiple target genes in plants over the past 2 years. But editing efficiency of the system in Arabidopsis is much lower compared with that in rice, because usually a vector containing CaMV 35S-drived Cas9 and a sgRNA Agrobacterium is introduced into plants by Agrobacterium-mediated floral dip, in which the embryo sac is the primary target of T-DNA. However, the CaMV 35S promoter has very low activity in Arabidopsis which leads to the low editing efficiency.

 

Researchers in Dr. XIE Qi’s group from the Institute of Genetics and Developmental Biology, the Chinese Academy of Sciences, reported a high genome editing efficiency CRISPR/Cas9 system. Their system could effectively improve gene mutation ratios in Arabidopsis.

 

Here, the Cas9 gene was driven by the promoter of YAO gene, which is preferentially expressed in tissues undergoing active cell division, and highly expressed in the embryo, embryo sac, endosperm and pollen. They introduced 35S:hSpCas9-BRI1-sgRNA or pYAO:hSpCas9-BRI1-sgRNA into the wild-type Arabidopsis to target BRI1 gene by Agrobacterium-mediated floral dip. pYAO:hSpCas9 generated a higher BRI1 mutation ratio (90.5%) compared to 35S:hSpCas9 (4.3%), and the T1 plants have more mutant alleles, some of which were heritable thought detection of Cas9-free T2 plants. PDS3 was selected to verify the system effect, the editing efficiency could reach 88.5%. This work provides a new CRISPR/Cas9 system to improve genome editing efficiency.

 

The study entitled “High efficiency genome editing in Arabidopsis using Yao promoter-driven CRISPR/Cas9 system” has been published in Molecular Plant (DOI: http://dx.doi.org/10.1016/j.molp.2015.10.004) with YAN Liuhua and WEI Shaowei as the co-first authors. This work was financially supported by grants from the National Natural Science Foundation of China and the grant 2014ZX08009-003-002. YAN Liuhua is supported by Chinese Postdoctoral Science Foundation.

 

Figure. pYAO:hSpCas9 edited Arabidopsis genes more efficiently than 35S:hSpCas9 through floral dip transformation. (Image by IGDB)