A CRISPR-Cas9 Vector System for Multiplex Targeting and Its Application in Editing Bentazon Resistance Gene CYP81A6 in Rice (Oryza sativa)

Journal Article / 2017

Nie Wang; Peng Wei; Yu Yaxin; Zhang Huan; Chen Qiusheng; Liu Changai; Liao Pengfei

The rice cytochrome P450 monooxgenase, which is encoded by CYP81A6 (Bel) gene, confers the resistance to bentazon and sulfonylurea herbicides. The recessive homozygous mutant (bel) of Bel shows bentazon sensitive lethality, which can be used as chemical lethal marker and will have important application values in hybrid rice seed production. In this study, two single guide RNAs were designed based on the first exon within the coding sequence of CYP81A6 gene. A total of 56 positive transgenic T0 plants were obtained, and the genomic DNA of 9 plants were extracted for testing the gene mutation by sequencing. The total gene mutation efficiency was 77.8%, and the homozygous mutation efficiency was up to 71.4%. After spraying bentazon, the homozygous mutants showed bentazon sensitive lethality. These results provided a theoretical basis to use CRISPR-Cas9 system for breeding bentazon-lethal male sterile lines and editing other function genes in rice.