Application of CRISPR/Cas2 System for Genome Editing in Plants

CRISPR/Cas system has developed a new technology to modify target genes. In this study, Cas Phi 2 is a newly Cas protein that we used for genome modification in Arabidopsis and tobacco. PDS and BRI1 of marker genes were chosen for targeting. Cas Phi 2 has the function to cleave pre-crRNA. In the presence of 10 mM Mg2+ irons concentration, sgRNA3 type guided Cas Phi 2 to edit target gene and generate mutation, and a mutant seedling of AtBRI1 gene with an expected male sterile phenotype was obtained. In the process of tobacco transformation, the gene editing activity of Cas Phi 2 can be activated by 100 nM Mg2+ irons concentration, and sgRNA1 type guided Cas Phi 2 to edit target gene. Mutant seedlings of NtPDS gene with an expected albino were obtained. The results indicate that Cas Phi 2 can effectively edit target genes under the guidance of different sgRNA type in the presence of Mg2+ ions. Together, our results verify that the CRISPR/Cas Phi 2 system is an effective and precise tool for genome editing in plants.