CRISPR/Cas9-mediated knockout of Bsr-d1 enhances blast resistance of rice in Northeast China

Abstract

Abstract Rice blast is a globaldisease, which has a significant negative impact on rice yield and quality. Due to the complexity and changeability of the physiological races of rice blast, controlling the rice blast is challenging in agricultural production. Bsr-d1, a negative transcription factor confers broad-spectrum resistance to rice blast which was identified from the indica rice cultivar Digu, however, the biological function in japonica rice varieties is still unclear. In this study, we analyzed the blast resistance allele of OsBsr-d1 in a total of 256 japonica rice varieties of Jilin province in Northeast China and found that it does not exist in these varieties. Therefore, we developed Bsr-d1 knockout mutants by CRISPR/Cas9 system using a japonica rice variety Jigeng88 (JG88) as a recipient variety. Compared with the wild-type JG88, the homozygous Bsr-d1 mutant lines, KO#1 and KO#2, showed enhanced leaf blast resistance at seedling stage to several Magnaporthe oryzae (M. oryzae) races collected from Jilin Province in Northeast China. Physiological and biochemical indicators revealed that the homozygous mutant lines produced more hydrogen peroxide compared to JG88 when infected by M. oryzae. Comparative RNA-seq revealed that the DEGs were mainly involved in the synthesis of amide compounds, zinc finger proteins and transmembrane transporters, etc. In summary, our results indicate that the Bsr-d1 knockout mutants developed through gene editing technology can enhance the broad-spectrum resistance of rice in Northeast China to rice blast. This study not only provides a theoretical basis for disease resistance breeding of Bsr-d1 gene in Northeast China, but also provides new germplasm resources for disease resistance rice breeding.