Rapid generation of genetic diversity by multiplex CRISPR/Cas9 genome editing in rice.

The clustered regularly interspaced short palindromic repeats (CRISPR)-associated endonuclease 9 (CRISPR/Cas9) system has emerged as a promising technology for specific genome editing in many species. Here we constructed one vector targeting eight agronomic genes in rice using the CRISPR/Cas9 multiplex genome editing system. By subsequent genetic transformation and DNA sequencing, we found that the eight target genes have high mutation efficiencies in the T0 generation. Both heterozygous and homozygous mutations of all editing genes were obtained in T0 plants. In addition, homozygous sextuple, septuple, and octuple mutants were identified. As the abundant genotypes in T0 transgenic plants, various phenotypes related to the editing genes were observed. The findings demonstrate the potential of the CRISPR/Cas9 system for rapid introduction of genetic diversity during crop breeding.