中国医科大学学报

中国医科大学学报
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中国医科大学学报 ›› 2018, Vol. 47 ›› Issue (6): 499-502.doi: 10.12007/j.issn.0258‐4646.2018.06.005

• 论著 • 上一篇    下一篇

MYCT1-GST原核表达载体的构建及鉴定

孙媛媛1, 刘佳2, 富伟能1   

  1. 1. 中国医科大学基础医学院医学遗传学教研室, 沈阳 110122;
    2. 中国医科大学实验动物部, 沈阳 110001
  • 收稿日期:2018-01-08 出版日期:2018-06-30 发布日期:2018-05-29
  • 通讯作者: 富伟能 E-mail:wnfu@cmu.edu.cn
  • 作者简介:孙媛媛(1991-),女,助理实验师,硕士.
  • 基金资助:
    国家自然科学基金(81372876);辽宁省自然科学基金(2015020518)

Construction and Identification of the MYCT1-GST Prokaryotic Expression Vector

SUN Yuanyuan1, LIU Jia2, FU Weineng1   

  1. 1. Department of Medical Genetic, College of Basic Medical Science, China Medical University, Shenyang 110122, China;
    2. The Ministry of Experimental Animals, China Medical University, Shenyang 110001, China
  • Received:2018-01-08 Online:2018-06-30 Published:2018-05-29

摘要: 目的 构建MYCT1-GST原核表达载体,并纯化MYCT1-GST融合蛋白。方法 以正常人外周血cDNA为模板,PCR方法扩增该基因的开放阅读框,定向克隆至pET28a (+)载体后送测序鉴定。将构建好的MYCT1-GST表达载体转化至BL21(DE3)菌株,通过IPTG诱导表达,PCR结合测序方法鉴定载体构建是否成功,Western blotting方法鉴定MYCT1-GST融合蛋白的表达。结果 测序结果证实成功将MYCT1的开放阅读框克隆至pET28a (+)表达载体,经诱导后成功纯化了MYCT1-GST融合蛋白。结论 成功构建MYCT1-GST表达载体,为进一步研究MYCT1相互作用蛋白质奠定了基础。

关键词: MYCT1, GST, 相互作用蛋白

Abstract: Objective To construct MYCT1-GST prokaryotic expression vector and purify MYCT1-GST recombinant protein. Methods Using cDNA synthesized from peripheral blood RNA as the template, the open reading frame of MYCT1 was PCR amplified and ligated into the pET28a(+) vector. The recombinant DNA was sequenced to verify successful vector construction. BL21(DE3) cells were transformed with the MYCT1-GST construct, and the expression of MYCT1-GST was induced by IPTG. Western blotting was performed to determine whether the recombinant protein was purified successfully. Results Sequencing and PCR results confirmed that the open reading frame of MYCT1 was cloned into pET28a(+) and its expression was induced by IPTG. Thus, we could successfully purify the MYCT1-GST recombinant protein. Conclusion We successfully designed the MYCT1-GST expression construct, which may aid further studies on proteins interacting with MYCT1.

Key words: MYCT1, GST, interaction protein

中图分类号: 

  • R394
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