Construction and identification of the RNA interference system targeting CGI-100 gene

WANG Yong-qiang; et al

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Construction and identification of the RNA interference system targeting CGI-100 gene

更新时间：2024-01-17

- Construction and identification of the RNA interference system targeting CGI-100 gene
- Journal of Chongqing Medical University Vol. 33, Issue 12, Pages: 1418-1422(2008)
- 作者机构：
  
  重庆医科大学临床生化教研室重庆医科大学临床检验诊断学教育部重点实验室
- 作者简介：
- 基金信息：
- DOI：
  CLC： R73-3
- Published：2008，
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[1]汪永强,张彦,雷燕.靶向CGI-100基因shRNA干扰载体的构建与鉴定[J].重庆医科大学学报,2008,33(12):1418-1422.

WANG Yong-qiang, et al. Construction and identification of the RNA interference system targeting CGI-100 gene. [J]. Journal of Chongqing Medical University 33(12):1418-1422(2008)
[1]汪永强,张彦,雷燕.靶向CGI-100基因shRNA干扰载体的构建与鉴定[J].重庆医科大学学报,2008,33(12):1418-1422. DOI：

WANG Yong-qiang, et al. Construction and identification of the RNA interference system targeting CGI-100 gene. [J]. Journal of Chongqing Medical University 33(12):1418-1422(2008) DOI：

摘要

目的:构建靶向CGI-100基因的RNAi表达载体并鉴定其抑制效率。方法:设计及化学合成3对靶向CGI-100基因的短发夹结构寡核苷酸

经退火成双链DNA片段

将其与经限制性内切酶BamHⅠ和HindⅢ双酶切的PGenesil-1质粒连接

构建表达shRNA的重组质粒

用脂质体2000转染到K562细胞中进行表达

经RT-PCR和斑点杂交检测转染前后CGI-100基因表达的变化。结果:经DNA测序证实3对靶向CGI-100基因的RNA干扰表达载体PGenesil-1-CGI-100构建成功

其中第1对PGenesil-1-CGI-100在mRNA水平抑制白血病K562细胞中CGI-100基因表达的效率最高

达54%。结论:成功构建针对CGI-100基因的RNA干扰的表达载体

并筛选出1对抑制效率较高的重组干扰载体

为进一步研究K562细胞中CGI-100基因的功能奠定基础。

Abstract

Objective:To construct the expression vector of RNA interference(RNAi)system for CGI-100 gene and identify its inhibition efficiency.Methods:Three oligonucleotides targeting CGI-100 gene were devised and chemically synthesized

annealed to form double-strand DNA and inserted into plasmid PGenesil-1 whose segments were digested by BamHⅠ and HindⅢ.The recombinant vector were then transfected into K562 cells with lipofectamineTM 2000.CGI-100 expresstion in the transfected cells was assayed by RT-PCR and dot blot.Results:The recombinant plasmids of PGenesil-1-CGI-100 were successfully constructed by DNA sequencing analysis.The first of PGenesil-1-CGI-100 had the highest inhibition efficiency in mRNA levels

up to 54%.Conclusion:The RNA interference system targeting CGI-100 gene were successfully constructed and a interference vector with a higher inhibition efficiency was screened

which provides the foundation for further investigation.

关键词

CGI-100基因RNA干扰shRNAK562细胞

Keywords

CGI-100 geneRNA interferenceshRNAK562 cell

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