设为首页  |  加入收藏  |  联系我们  |  English  |  中国科学院  
 首  页  实验室概况 研究队伍 科研工作 平台建设 开放交流 人才培养 规章制度
 
  当前位置:首页 > 最新论文  
Genotyping Genome‐edited Mutations in Plants Using CRISPR Ribonucleoprotein Complexes.
时间:2018-05-24 来源: 点击:

Genotyping Genome‐edited Mutations in Plants Using CRISPR Ribonucleoprotein Complexes

Zhen Liang, Kunling Chen, Yan Yan, Yi Zhang, Caixia Gao


Plant Biotechnology Journal
DOI:10.1111/pbi.12938

Abstract

 

Despite the great achievements in genome editing, accurately detecting mutations induced by sequence‐specific nucleases is still a challenge in plants, especially in polyploidy plants. An efficient detection method is particularly vital when the mutation frequency is low or when a large population needs to be screened. Here, we applied purified CRISPR ribonucleoprotein complexes to cleave PCR products for genome‐edited mutation detection in hexaploid wheat and diploid rice. We show that this mutation detection method is more sensitive than Sanger sequencing and more applicable than PCR/RE method without the requirement for restriction enzyme site. We also demonstrate that this detection method is especially useful for genome editing in wheat, because target sites are often surrounded by single nucleotide polymorphisms. By using this screening method, we were also able to detect foreign DNA‐free tagw2mutations induced by purified TALEN protein. Finally, we show that partial base editing mutations can also be detected using high fidelity SpCas9 variants or FnCpf1. The PCR/RNP method is low‐cost and widely applicable for rapid detection of genome‐edited mutation in plants.
 
【打印本页】【关闭本页】
Copyright © 植物细胞与染色体工程国家重点实验室(中国科学院遗传与发育生物学研究所)
地址:北京市朝阳区北辰西路1号院2号 邮编:100101 电话: 010-64806537 传真: 010-64806537