中国水稻科学2016，Vol.30Issue(5)：469-478,10.DOI:10.16819/j.1001-7216.2016.6009
利用 CRISPR/Cas9技术敲除水稻 Pi21基因的效率分析
Knock-out Efficiency Analysis of Pi21 Gene Using CRISPR/Cas9 in Rice
摘要
Abstract
A CRISPR/Cas9 vector,which contains two targets (Target 1 and Target 2)of Pi21 gene,was constructed and transformed to rice variety Nanjing 9 108.Twenty-eight T0 transgenic lines were obtained and confirmed by T-DNA specific PCR.Restriction digestion analysis revealed 78.57% mutant ratio in Target 1 site,92.86% mutant ratio in Target 2 site,78.57% ratio of lines containing both Target 1 and Target 2 mutant in transgenic lines.Through sequencing analysis,we found the mutant types of targets,such as deletion of bases,insertion of bases,insertion behinds deletion of bases,and long fragment deletion of DNA,etc.Moreover,except amino acid deletion,most transgenic lines displayed frameshift mutation,which caused function loss of Pi21 .In this way,a series of Pi21 -knock-out rice lines were obtained,and further,could be used for functional analysis of Pi21 and rice blast broad-spectrum resistant breeding.关键词
CRISPR/Cas9/稻瘟病/Pi21/基因敲除Key words
CRISPR/Cas9/rice blast/Pi21/gene knock-out分类
生物科学引用本文复制引用
王芳权,范方军,李文奇,朱金燕,王军,仲维功,杨杰..利用 CRISPR/Cas9技术敲除水稻 Pi21基因的效率分析[J].中国水稻科学,2016,30(5):469-478,10.基金项目
江苏省现代农业重点研发项目(BE2015355) （BE2015355）
国家重大农技推广项目(NG[15]003) （NG[15]003）
国家自然科学基金资助项目(31301652) （31301652）
江苏省自然科学基金资助项目(BK20130723) （BK20130723）
