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Cloning and Expression of Human sTNFR1 in E.Coli JM109

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Author:
No author available
Journal Title:
JOURNAL OF CHINESE PHYSICIAN
Issue:
7
DOI:
10.3760/cma.j.issn.1008-1372.2005.07.002
Key Word:
人可溶性肿瘤病坏死因子受体1;克隆;原核表达

Abstract: 目的构建人可溶性肿瘤坏死因子受体1基因的重组质粒,进行原核表达.方法以Hela细胞的总RNA为模板,用RT-PCR方法扩增sTNFR1全编码区基因片段,构建含有目的片段的T载体克隆及原核表达载体pMAL-c2x重组质粒亚克隆,经异丙基-β-D半乳糖苷酶(IPTG)诱导重组质粒菌表达sTNFR1,以淀粉树脂亲和层析法纯化重组蛋白,用十二烷基硫酸钠聚丙烯酰胺凝胶电泳(SDS-PAGE)对重组蛋白进行分析.结果经核苷酸序列测序和SDS-PAGE法鉴定,成功构建了人sTNFR1重组质粒基因工程菌,并表达和纯化了sTNFR-MBP融合蛋白.结论成功地构建了人sTNFR1重组质粒克隆,表达并纯化了sTNFR1-MBP融合蛋白.

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