Genome editing with miniature size Cas nuclease; SpCas12f in Oryza sativa

In genome editing using the CRISPR-Cas system, Cas9 derived from Streptococcus pyogenes (SpCas9) is widely used due to its high activity, but it has a large molecular weight (1368 amino acid residues). Just recent year, targeted mutagenesis in human cells and maize using Cas12f derived from Syntrophomonas palmitatica (SpCas12f), a very small Cas with 497 amino acid residues which is more suitable size for virus vectors, was reported. However, there are no reports of genome editing using SpCas12f in other crops. In this study, we applied SpCas12f to genome editing in rice, which is one of the most important staple crops in the world. SpCas12f and gRNA including target sequence of OsTubulin were introduced into rice calli, and mutation introduction was confirmed.