贵州医科大学学报

2004, (02) 99-101+104

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G156A六氧甲基鸟嘌呤-DNA-甲基转移酶原核表达载体的构建和表达
The Construction and Expression of Prokaryotic Expression Vector of G156A O~6-methylguanine-DNA-methyltransferase

肖芸,王季石,孙等军,任志娟,张维,吴正宇,张旭,方琴
XIAO Yun 1, WANG Ji-shi 2, SUN Deng-jun 3, REN Zhi-juan 2, ZHANG Wei 2, WU Zheng-yu 3, ZHANG Xu 2, FANG Qin 2 (1. Department of Clinical Hematological Examinations, The Affiliated Hospital of Guiyang Medical College, Guiyang 550004, China; 2. Department of Hematology, The Affiliated Hosptial of Guiyang Medical College, Guiyang 550004, China; 3. Department of Pharmaceutics, The Affiliated Hosptial of Guiyang Medical College, Guiyang 550004, China)

摘要(Abstract):

目的 :构建原核表达载体pPROEXHTb G15 6AMGMT ,观察突变型G15 6AMGMT在大肠杆菌中的表达情况。方法 :通过基因重组技术将G15 6AMGMT亚克隆至pPROEXHTb原核表达载体 ,在大肠杆菌DH5α中经异丙基硫代 β D半乳糖苷 (IPTG)诱导表达。 结果 :经PCR及酶切分析证实 ,成功构建了pPROEXHTb G15 6AMGMT原核表达载体 ,经SDS 聚丙烯酰胺凝胶电泳 (SDS PAGE)显示重组克隆化基因pPROEXHTb G15 6AMGMT在大肠杆菌DH5α中有表达。结论 :G15 6AMGMT在大肠杆菌DH5α中有表达 ,为获取突变型MGMT工程蛋白奠定了基础。
Objective: To construct recombinant prokaryotic expression vector pPROEX HTb-G156A MGMT and to observe its expression in E.coli DH5α. Methods: The G156 MGMT cDNA was subcloned into pPROEX HTb. G156A MGMT protein was induced by IPTG to express in E.coli. Results: The recombinant vector, pPROEX HTb-G156A MGMT, was successfully constructed and confirmed by the restriction endonuclease and PCR analysis. G156A MGMT protein expressed successflly in E.coli. Conclusions: G156A MGMT expresses in E.coli under the induction of IPTG. The study provides a foundation to the research aimed at obtaining mutant protein of G156A MGMT.

关键词(KeyWords): 六氧甲基鸟嘌呤-DNA-甲基转移酶;大肠杆菌;基因表达
O 6-methylguanine-DNA-methyltransferase; Escherichia coli; gene expression

Abstract:

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基金项目(Foundation): 贵州省科委基金资助项目(D2 0 0 0 - 5)

作者(Author): 肖芸,王季石,孙等军,任志娟,张维,吴正宇,张旭,方琴
XIAO Yun 1, WANG Ji-shi 2, SUN Deng-jun 3, REN Zhi-juan 2, ZHANG Wei 2, WU Zheng-yu 3, ZHANG Xu 2, FANG Qin 2 (1. Department of Clinical Hematological Examinations, The Affiliated Hospital of Guiyang Medical College, Guiyang 550004, China; 2. Department of Hematology, The Affiliated Hosptial of Guiyang Medical College, Guiyang 550004, China; 3. Department of Pharmaceutics, The Affiliated Hosptial of Guiyang Medical College, Guiyang 550004, China)

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DOI: 10.19367/j.cnki.1000-2707.2004.02.003

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