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[1]董 旭,张兆域,苟芳琳,等.利用CRISPR/Cas9 系统构建稳定敲除CBX2基因的A549细胞系[J].天津医科大学学报,2018,24(05):395-398.
 DONG Xu,ZHANG Zhao-yu,GOU Fang-lin,et al.Construction of CBX2 gene knockout stable A549 cell line with CRISPR /Cas9 system[J].Journal of Tianjin Medical University,2018,24(05):395-398.
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《天津医科大学学报》[ISSN:1006-8147/CN:12-1259/R]

卷:
24
期数:
2018年05期
页码:
395-398
栏目:
基础医学
出版日期:
2018-09-20

文章信息/Info

Title:
Construction of CBX2 gene knockout stable A549 cell line with CRISPR /Cas9 system
作者:
董 旭张兆域苟芳琳兰 蓓
天津医科大学生物化学与分子生物学系,天津 300070
Author(s):
DONG Xu ZHANG Zhao-yu GOU Fang-lin LAN Bei
Department of Biochemistry and Molecular Biology, Tianjin Medical University, Tianjin 300070, China
关键词:
CBX2基因CRISPR/Cas9 系统基因敲除
Keywords:
CBX2 gene CRISPR/Cas9 system gene knockout
分类号:
Q7
DOI:
-
文献标志码:
A
摘要:
目的:利用CRISPR/Cas9系统构建稳定敲除CBX2基因的A549细胞系。方法:设计针对敲除CBX2的short guide RNA(sgRNA),并克隆到载体PX459中。将测序正确的重组质粒转染到A549细胞中,利用嘌呤霉素筛选转染阳性细胞并分离得到5个单克隆细胞系,通过Western blot方法检测构建的细胞系中CBX2蛋白表达情况。结果:成功构建了靶向CBX2的CRISPR/Cas9重组质粒,筛选出的5个单克隆细胞系中CBX2蛋白表达水平均显著下降。结论:成功利用CRISPR/Cas9 系统构建了稳定敲除CBX2基因的A549细胞系。
Abstract:
Objective: To construct CBX2 gene knockout stable A549 cell line with CRISPR/Cas9 system. Methods:The single guide RNA (sgRNA) targeting CBX2 gene was designed and inserted into the expression vector PX459 using molecular cloning technique. The recombined plasmid was verified by sequencing. The plasmid was transfected into A549 cell line. After puromycine selection, monoclonal cell line was picked for expansion. Finally, the expression of CBX2 protein in A549 CBX2 knockout cell line was detected by Western blot. Results:The CRISPR / Cas9 recombinant plasmid targeting CBX2 was successfully constructed, and the expression of CBX2 protein was significantly decreased. Conclusion:The A549 CBX2 knockout cell line could be successfully constructed by CRISPR/Cas9 system.

参考文献/References:


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更新日期/Last Update: 2018-09-30