Construction and identification of the chimeric mtb8.4/hIL12  eukaryotic expression plasmid

LI Hui; et al

doi:10.13406/j.cnki.cyxb.2004.05.005

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Construction and identification of the chimeric mtb8.4/hIL12 eukaryotic expression plasmid

更新时间：2024-01-17

- Construction and identification of the chimeric mtb8.4/hIL12 eukaryotic expression plasmid
- Journal of Chongqing Medical University Issue 5, Pages: 576-579(2004)
- 作者机构：
  
  1. 重庆医科大学第二临床学院感染科
  2. 泸州医学院附属医院感染病科
  3. 重庆医科大学第二临床学院感染科,重庆,400010
  4. ,四川,泸州,646000
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.2004.05.005
  CLC： R346
- Published：2004，
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[1]李晖,任小华,钟森,任红.结核分枝杆菌mtb8.4/hIL12嵌合基因的克隆及真核表达质粒的构建[J].重庆医科大学学报,2004(05):576-579+587.

LI Hui, et al. Construction and identification of the chimeric mtb8.4/hIL12 eukaryotic expression plasmid. [J]. Journal of Chongqing Medical University (5):576-579(2004)
[1]李晖,任小华,钟森,任红.结核分枝杆菌mtb8.4/hIL12嵌合基因的克隆及真核表达质粒的构建[J].重庆医科大学学报,2004(05):576-579+587. DOI： 10.13406/j.cnki.cyxb.2004.05.005.

LI Hui, et al. Construction and identification of the chimeric mtb8.4/hIL12 eukaryotic expression plasmid. [J]. Journal of Chongqing Medical University (5):576-579(2004) DOI： 10.13406/j.cnki.cyxb.2004.05.005.

摘要

目的 :克隆结核分枝杆菌mtb8.4 /hIL12嵌合基因

构建其真核表达质粒

并进行鉴定。方法 :采用基因工程技术

首先以 pcDNA 3.1（+） -mtb 8.4为模板

经聚合酶链反应（PCR）扩增出mtb 8.4 -linker基因

与pCI-neo载体进行连接重组

构建成pCI-neo -mtb 8.4 -linker（pML）重组质粒

然后以pORF -hIL12质粒为模板

经PCR扩增出hIL - 12基因

将hIL -12基因与pML质粒进行连接重组

构建成mtb 8.4 /hIL12嵌合基因真核表达质粒

用限制性内切酶消化、PCR及DNA序列测定等多种分子生物学方法进行鉴定。结果 :mtb 8.4 /hIL12嵌合基因重组真核表达质粒经证实构建成功。结论 :mtb 8.4 /hIL12嵌合基因重组真核表达质粒的成功构建

为进一步研究其免疫保护效果及制备结核病mtb 8.4 /hIL12嵌合基因疫苗奠定了基础

Abstract

Objective:To construct and identify the chimeric mtb8.4/hIL12 eukaryotic expression plasmid.Methods:Firstly mtb8.4-linker was amplified by PCR and cloned into the single Nhe Ⅰ and Mlu Ⅰ cloning sites of pCI-neo

so pCI-neo-mtb8.4-linker(pML) plasmid was constructed.Secondly hIL-12 was amplified by PCR and cloned into the single Mlu Ⅰ and Sal Ⅰ cloning sites of pML.Finally correct pCI-neo-mtb8.4/hIL12 (pMI) plasmid was identified by PCR

RE digestion and DNA seguencing.Results:The accuracy of pMI plasmid construction was confirmed by a number of molecular biological techniques.Conclusion:The construction of mtb8.4/hIL12 chimera by linkage of M.tuberculosis mtb8.4 gene to human IL12 gene provides the possibility for investigating a new tuberculosis vaccine.

关键词

结核病Mtb8.4hIL-12嵌合基因

Keywords

tuberculosisMtb8.4hlL-12Chimeric gene

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