生物学杂志

生物学杂志 ›› 2021, Vol. 38 ›› Issue (2): 43-.doi: 10.3969/j.issn.2095-1736.2021.02.043

• 研究报告 • 上一篇    下一篇

MAR序列对逆转录病毒载体表达外源基因的调控作用

  

  1. 新乡医学院三全学院，新乡453003
  • 出版日期:2021-04-18 发布日期:2021-04-25
  • 作者简介:张　靖，硕士，讲师，研究方向为真核基因表达调控与基因工程， E⁃mail：739353441@qq.com
  • 基金资助:
    国家自然科学基金（NSFC-河南联合基金项目）（编号U1604193）

Regulation of MAR⁃mediated retroviral vector on transgene expression

  1. Sanquan College of Xinxiang Medical University，Xinxiang 453003，China
  • Online:2021-04-18 Published:2021-04-25

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摘要： 为研究核基质结合区对逆转录病毒载体表达外源基因的调控作用，将CAT基因插入到载体pLXSN中构建出中间载体，构建出在5'端和3'端含人β⁃珠蛋白核基质结合区的载体。逆转录载体转染PA317细胞后G418筛选，提取PA317阳性细胞病毒上清液转染Bel⁃7404，经G418筛选出阳性细胞。测定3组CAT报告基因表达量。结果显示，与中间载体相比，3'端含核基质结合区的载体可以显著提高CAT报告基因的表达量（P< 0.05），而5'端含MAR序列的载体不能显著提高基因表达水平（P> 0.05）。以上结果表明MAR介导逆转录表达载体能提高外源报告基因CAT的表达水平，且MAR对转基因表达调控作用与其在载体上插入位置有关。

关键词: 核基质结合区, 基因沉默, 基因治疗, 位置效应

Abstract: In order to research the regulation of MAR⁃mediated retroviral vector on transgene expression，the report geneCATwas firstly inserted into retroviral vector，the cloned β⁃globin MAR DNA fragment was then inserted respectively into 5'or 3'ends of the vectors，resulting the retroviral vectors series contained MAR in different sites. The recombinated vectors were transfected into PA317 cells. The positive cells were screened by G418，Bel⁃7404 was screened by G418 after transfected by virus from PA317, and theCATgene expression was analyzed by ELISA. The results showed that，compared to the control，the MAR inserted at the 3' end of the vector could enhance theCATgene expression significantly by 11 times（P<0.05），while theCATgene expression was not enhanced when the MAR was inserted at the 5' end of the vector（P>0.05）. These results indicated that MAR could increase the expression level of reporter gene and the regulation of MAR on transgene expression is relied on the inserted site.

Key words: matrix attachment region, gene silencing, gene therapy, position effect

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