Development of high-quality fragrant indica CMS line by editing Badh2 gene using CRISPR-Cas9 technology in rice（Oryza sativa L.）

Fragrance is one of the important traits for quality improvement in rice, which is mainly controlled by the Badh2 gene encoding a betaine aldehyde dehydrogenase on chromosome 8. The mutation of Badh2 gene can increase the content of 2-acetyl-1-pyrroline（2-AP） and promote the fragrance production in rice. In this study, the Badh2 gene in Mingtai B（MTB）, an elite maintainer line of Indica hybrid rice showing high eating quality bred by Sanming Academy of Agricultural Sciences, was edited and knocked out by using CRISPR-Cas9 technology. Two T0 transgenic lines carrying homozygous mutation on the loci of Badh2 were generated, and 48 T1 individuals derived from these two plants were genotyped by PCR amplification and sequencing analysis. A mutant line named MTB-badh2, which had a single T nucleotide insertion in the second exon of Badh2 without the vector skeleton, was finally obtained. In our study, the expression of Badh2 and the content of 2-AP were determined by semi-quantitative RT-PCR, qRT-PCR, and gas chrom