靶向呼吸道合胞病毒M2基因pshRNA载体质粒的构建及意义

崔玉霞; 王萍玲; 周娟; 杨锡强

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靶向呼吸道合胞病毒M2基因pshRNA载体质粒的构建及意义

更新时间：2024-01-17

- 靶向呼吸道合胞病毒M2基因pshRNA载体质粒的构建及意义
- Construction and significance of the short hairpin RNA recombinant plasmid targeting M2 gene of respiratory syncystial virus
- 重庆医科大学学报 2006年第1期页码：1-4
- 作者机构：
  
  1. 重庆医科大学附属儿童医院免疫室
  2. 贵阳市妇幼保健医院妇产科
  3. 重庆医科大学附属儿童医院免疫室,重庆,400014
  4. ,贵阳,550004
- 作者简介：
- 基金信息：
  
  国家自然科学基金资助项目（30340045）
- DOI：
  中图分类号： R373
- 纸质出版日期：2006，
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[1]崔玉霞,王萍玲,周娟,杨锡强.靶向呼吸道合胞病毒M2基因pshRNA载体质粒的构建及意义[J].重庆医科大学学报,2006(01):1-4.

CUI Yuxia, et al. Construction and significance of the short hairpin RNA recombinant plasmid targeting M2 gene of respiratory syncystial virus[J]. Journal of Chongqing Medical University, 2006,(1):1-4.
[1]崔玉霞,王萍玲,周娟,杨锡强.靶向呼吸道合胞病毒M2基因pshRNA载体质粒的构建及意义[J].重庆医科大学学报,2006(01):1-4. DOI：

CUI Yuxia, et al. Construction and significance of the short hairpin RNA recombinant plasmid targeting M2 gene of respiratory syncystial virus[J]. Journal of Chongqing Medical University, 2006,(1):1-4. DOI：

摘要

目的:构建针对呼吸道合胞病毒（RSV）M2基因m RNA的短发卡结构状RNA（shRNA）重组载体质粒

为利用RNA干扰技术抗RSV感染的深入研究奠定基础;方法:按shRNA设计原则

选择RSV-M 2基因作为靶基因

设计19bp长的能产生短发卡结构的两段DNA反向重复序列

中间以9bp序列间隔

经退火形成互补双链

克隆至转录载体pgenesil-1上

重组质粒经酶切和测序鉴定

然后将构建成功的重组质粒转染H EP2细胞

荧光显微镜下观察绿色荧光细胞发生率以评估转染效率。结果:将设计合成的DNA片段成功克隆至载体上

经酶切及序列鉴定为目的序列

并且荧光显微镜下观察细胞有较多的绿色荧光蛋白表达。结论:成功构建了针对RSV M 2基因m RNA的shRNA重组载体质粒

可利用重组质粒进一步研究其对RSV感染的抑制

从而为抑制RSV感染寻找新的基因治疗手段。

Abstract

Objective: To construct the recombinant plasmid targeting M2 gene of respiratory syncystial virus(RSV) for further study of RNA interference(RNAi) technology in inhibiting RSV infection.Methods: According to the designing principles of shRNA

The M2 gene of RSV was selected as the target gene.A 19bp reverse repeated sequences with 9bp spacer were designed and synthesized.The complement strain were obtained by annealing and inserted into the vector pgenesil-1 containing green fluorescenet protein(GFP) sequence and U6 promotor.Finally the recombinant plasmid were identified by enzyme digesion and DNA sequencing

then transfected into HEP2 cells and observed by fluorescence microscope.Results: The recombinante plasmid targeting the mRNA of RSV M2 gene was successfully constructed and the good efficents of transfection were found by fluorescence microscope.Conclusion: The successful construction of the pshRNA recombinant plasmid targeting the M2 gene of RSV will help futher study on application of the RNAi technology to anti-RSV infection.

关键词

呼吸道合胞病毒M2基因短发卡结构状RNA重组质粒

Keywords

Respiratory syncystial virusM2 geneShort hairpin RNArecombinant plasmid

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