文章摘要
高逸湘,张评浒.人源细胞色素P450 2E1基因的克隆及其在大肠杆菌中的表达[J].南京医科大学学报,2010,(11):1542~1545
人源细胞色素P450 2E1基因的克隆及其在大肠杆菌中的表达
Cloning of human cytochrome P450 2E1 from Chinese and its expression in Escherichia coli
投稿时间:2010-06-10  
DOI:10.7655
中文关键词: 细胞色素P450 2E1  突变  表达
英文关键词: cytochrome P450 2E1  mutation  expression
基金项目:新药筛选核心技术规范化平台建设及其科学应用(2009ZX09302)
作者单位
高逸湘 南京医科大学第一附属医院麻醉科,江苏 南京 210029 
张评浒 中国药科大学新药筛选中心,江苏 南京 210009 
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中文摘要:
      目的: 人源细胞色素P450 2E1(cytochrome P450 2E1, CYP2E1)基因的克隆及其表达?方法:采用RT-PCR方法对我国汉族人CYP2E1基因进行了克隆,并利用pET-32a(+)原核表达载体进行了表达?结果:经过酶切与测序鉴定表明扩增的CYP2E1基因含有完整的编码序列,除该基因编码区第1 263位核苷酸发生C→T的突变(未引起氨基酸发生突变)外,其他核苷酸与氨基酸序列与GenBank中登录发表的人CYP2E1基因完全相同?SDS-PAGE分析结果显示该基因片段连接到原核表达载体pET-32a(+)后在BL21(DE3)大肠杆菌中成功表达?结论:该基因的成功表达为开展临床前药物安全性评价与药物代谢分析的后续工作打下了基础?
英文摘要:
      Objective: To clone and express the human cytochrome P450 2E1(CYP2E1) gene. Methods: Using RT-PCR, a full length gene encoding CYP2E1 was successfully cloned and its expression was analyzed in the host cell E. coli BL21(DE3)by SDS-PAGE. Results: Except a C→T mutation at 1 263 nucleotide site, the nucleotide sequences of this CYP2E1 gene are consistent with the sequences in GenBank. The CYP2E1 gene was successfully inserted into the expression vector pET-32a (+) and expressed in the host cell E. coli BL21(DE3). The CYP2E1 protein with relative molecular mass of approximately 67ku was observed on the SDS-PAGE gel. Conclusion:The cloning and expression of CYP2E1 provide the foundation for following pre-clinical drug safety evaluation and the analysis of drug metabolism.
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