| 常会波1 朱彦丽2 王立文2 李尔珍2 刘卓1 欧阳胜荣1 吴建新1△.甲基丙二酰辅酶A 变位酶表达载体的构建及初步表达[J].,2012,12(13):2414-2417 |
| 甲基丙二酰辅酶A 变位酶表达载体的构建及初步表达 |
| The Construction and Expression of Methymalonyl-CoA Mutase（MCM）Expressive Vector |
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| DOI： |
| 中文关键词: 甲基丙二酰辅酶A 变位酶（MCM） MUT 基因 表达 |
| 英文关键词: Methymalonyl-CoA mutase( MCM) MUT gene Expression |
| 基金项目:国家重点基础研究发展计划(2007CB511903)；北京市优秀人才资助项目（20081D0303200106）；
首都特色临床医学技术发展研究（D101100050010040） |
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| 中文摘要: |
| 目的：克隆表达甲基丙二酰辅酶A 变位酶(MCM)蛋白,为进一步研究相关基因突变对功能的影响机制奠定基础。方法：自人
外周血淋巴细胞中提取总RNA、逆转录,并与pET32a 构建融合蛋白原核表达载体;优化蛋白表达诱导条件; 经SDS-PAGE、Western
Blot 检测目的蛋白的表达。结果：经酶切鉴定并经测序证实获得全长2210bp 的甲基丙二酰辅酶A 变位酶基因（MUT）,并成功构
建融合蛋白原核表达载体,SDS-PAGE 在102 kDa 处获得目的条带,Western Blot 检测确定为MCM 表达蛋白。结论：成功克隆表达
出MCM 表达蛋白。 |
| 英文摘要: |
| Objective: To clone and express MUT gene in order to study the function and mechanism of MUT gene, which lay the
foundation for further study of mutations mechanisms. Methods: RNA was extracted from human lymphocytes and transcribed reversely,
and MUT gene was inserted into pET32a to construct fusion protein prokaryotic expression vector. The condition for induction and expression
was optimized. The fusion protein was detected by SDS-PAGE and Western Blot. Results: MUT gene, of 2210bp, was obtained
and identified by sequencing. MCM fusion protein prokaryotic expression vector were constructed successfully. A protein band of about
102 kDa was detected by SDS-PAGE, and the protein was verified to be MCM by Western-Blot. Conclusion: MUT gene was cloned and
expressed successfully. |
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