利用CRISPR/Cas9基因编辑技术改良大粒香稻瘟病抗性

Improvement of Blast Resistance of Rice Variety Dalixiang by CRISPR/Cas 9Gene Editing Technology

优质稻米大粒香是采用粳籼杂交方法育成的香稻品系,多年来是贵州省优质稻米企业开发生产的主推品种。但稻瘟病抗性差,限制了其大规模推广应用。本研究以大粒香为材料,根据CRISPR/Cas 9靶点设计要求设计稻瘟病基因pid3特异性靶点sgRNA序列,构建crispr-pid3双靶点载体,通过农杆菌介导遗传转化大粒香愈伤组织,获得45株转基因苗,有30株阳性苗。在T1代中共获得9株无Cas 9转基因位点的pid3纯合突变体,对其进行稻瘟病病情鉴定,与原种大粒香相比,稻瘟病抗性均有一定程度提高。且对改良株系进行米质鉴定,与野生型大粒香相比,稻米品质并未发生显著改变。说明改良株系的米质并未因pid3基因的编辑而受到影响。

High-quality rice variety Dalixiang belongs to aromatic rice line by hybrid between japonica and indica,which has been the main variety of high quality rice in Guizhou province for many years.However,its large-scale application is limited due to the poor resistance of rice blast.In this study,the dual target vector crispr-pid 3 was constructed according to specific target sgRNA sequence of Pid3 gene of rice blast based on the requirements of CRISPR/Cas 9 target design,and the callus of the Dalixiang was transformed by crispr-pid 3-contained Agrobacterium tumefaciens.Finally,a total of 45 transgenic seedlings were obtained,A total of 9 homozygous pid3 mutants without Cas 9-transgenic locus were obtained in T1 generation,and the disease conditions of these mutants were also identified.Compared with the original seed,the resistance of pid3 mutants to rice blast was improved to a certain extent.The quality identification of the improved rice lines showed that the quality of mutants was not significantly changed compared with the wild type,which these results indicated that the rice quality of the modified line was not affected by pid3 gene editing.