|本期目录/Table of Contents|

[1]刘郁莹,崔晓腾,高星杰,等.利用改良的CRISPR/Cas9基因编辑系统构建HeLa细胞SND1基因敲除稳定株[J].天津医科大学学报,2015,21(06):480-483.
　LIU Yu-ying,CUI Xiao-teng,GAO Xing-jie,et al.Construction of HeLa SND1 knockout gene stable strain by using modified CRISPR/Cas9 gene editing system[J].Journal of Tianjin Medical University,2015,21(06):480-483.
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利用改良的CRISPR/Cas9基因编辑系统构建HeLa细胞SND1基因敲除稳定株(PDF)
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《天津医科大学学报》[ISSN:1006-8147/CN:12-1259/R]

卷:
21卷
期数:
2015年06期
页码:
480-483
栏目:
基础医学
出版日期:
2015-11-20

文章信息/Info

Title:
Construction of HeLa SND1 knockout gene stable strain by using modified CRISPR/Cas9 gene editing system
文章编号:
1006-8147（2015）06-0480-04
作者:
刘郁莹1; 崔晓腾1; 高星杰2; 赵秀娟3; 付 雪1; 葛 林1; 苏 超2; 杨 洁1; 2
(天津医科大学1.生物化学系;2.基础医学研究中心;3.细胞生物学系,天津 300070)
Author(s):
LIU Yu-ying1;  CUI Xiao-teng1;  GAO Xing-jie2;  ZHAO Xiu-juan3;  FU Xue1;  GE Lin1;  SU Chao2;  YANG Jie1.2
(Tianjin Medical University 1.Department of Biochemistry; 2. Research Center of Basic Medical Science; 3. Department of Cell Biology, Tianjin 300070, China )
关键词:
SND1; 基因敲除; CRISPR/Cas9; HeLa 细胞
Keywords:
SND1;  gene knockout;  CRISPR/Cas9;  HeLa cells
分类号:
Q7
DOI:
-
文献标志码:
A
摘要:
目的：利用改良的CRISPR/Cas9基因编辑系统敲除HeLa细胞中SND1基因，构建HeLa细胞SND1基因敲除稳定株。方法：设计一对特异性识别SND1基因第二个启动子的上下游sgRNA，以PX462质粒为载体，构建出一对重组真核表达质粒。酶切和测序鉴定后，将一对重组质粒共同转染进入HeLa细胞中，使用嘌呤霉素进行阳性细胞筛选，挑取单克隆细胞进行培养。最后用Western Blot鉴定敲除效果。结果：sgRNA正确插入到PX462质粒载体中，转染并筛选单克隆后的细胞中没有SND1蛋白的表达。结论：成功构建出HeLa细胞SND1基因敲除稳定株。
Abstract:
Objective: To Apply modified CRISPR/Cas9 gene editing system to knock out the SND1 gene in HeLa cell and construct HeLa SND1 gene knockout stable strain. Methods: A pair of sgRNAs that could specially identify the upstream and downstream of SND1 gene second promoter was designed, then a recombinant eukaryotic expressional plasmid by the carrier of PX462 was constructed. After enzyme digestion and sequencing, a pair of recombinant plasmids into HeLa cell was co-transfected, then puromycin was used to screen positive cell and the monoclonal cell was developed. The knockout effect was measured by western blotting. Results: sgRNA was correctly inserted into the PX462 recombinant plasmid, and no SND1 protein was detected in HeLa cell after transfection and screening of monoclonal cell. Conclusion: HeLa SND1 gene knockout stable strain can be successfully built

参考文献/References:

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备注/Memo

备注/Memo:

?作者简介 刘郁莹（1986- ），女，硕士在读，研究方向：肿瘤免疫；

通信作者：杨洁，Email：yangji@tmu.edu.cn。



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更新日期/Last Update: 2015-11-27