一种用于筛选CRISPR/Cas9诱导的纯合/双等位基因突变体的简单且经济高效的方法。
A simple and cost-effective method for screening of CRISPR/Cas9-induced homozygous/biallelic mutants.
作者信息
Guo Jinggong, Li Kun, Jin Lifeng, Xu Rui, Miao Kaiting, Yang Fengbo, Qi Chaoya, Zhang Lin, Botella Jose R, Wang Ran, Miao Yuchen
机构信息
1State Key Laboratory of Cotton Biology, Department of Biology, Institute of Plant Stress Biology, Henan University, 85 Minglun Street, Kaifeng, 475001 China.
Zhengzhou Tabacco Research Institute of CNTC, No. 2 Fengyang Street, Zhengzhou, 450001 Henan China.
出版信息
Plant Methods. 2018 May 29;14:40. doi: 10.1186/s13007-018-0305-8. eCollection 2018.
BACKGROUND
The CRISPR/Cas9 system is being used for genome editing purposes by many research groups in multiple plant species. Traditional sequencing methods to identify homozygous mutants are time-consuming, laborious and expensive.
RESULTS
We have developed a method to screen CRISPR/Cas9-induced mutants through Mutation Sites Based Specific Primers Polymerase Chain Reaction (MSBSP-PCR). The MSBSP-PCR method was successfully used to identify homozygous/biallelic mutants in and , and we speculate that it can be used for the identification of CRISPR/Cas9-induced mutants in other plant species. Compared to traditional sequencing methods, MSBSP-PCR is simpler, faster and cheaper.
CONCLUSIONS
The MSBSP-PCR method is simple to implement and can save time and cost in the screening of CRISPR/Cas9-induced homozygous/biallelic mutants.
背景
许多研究小组正在多种植物物种中使用CRISPR/Cas9系统进行基因组编辑。传统的用于鉴定纯合突变体的测序方法既耗时、费力又昂贵。
结果
我们开发了一种通过基于突变位点的特异性引物聚合酶链反应（MSBSP-PCR）筛选CRISPR/Cas9诱导突变体的方法。MSBSP-PCR方法已成功用于鉴定[具体物种1]和[具体物种2]中的纯合/双等位基因突变体，并且我们推测它可用于鉴定其他植物物种中CRISPR/Cas9诱导的突变体。与传统测序方法相比，MSBSP-PCR更简单、快速且成本更低。
结论
MSBSP-PCR方法易于实施，并且在筛选CRISPR/Cas9诱导的纯合/双等位基因突变体时可节省时间和成本。
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