人可溶性肿瘤坏死因子受体1基因真核表达载体的构建

Construction of eukaryotic vector of human sTNFR1

目的构建人可溶性肿瘤坏死因子受体1基因(sTNFR1)的真核表达载体,为进一步研究打下基础。方法以Hela细胞的总RNA为模板,用RT-PCR方法扩增人sTNFR1全编码区基因片段,构建含有目的片段的T载体克隆及真核表达载体pCDNA3.1(-)重组质粒亚克隆,并用酶切分析和序列测定进行鉴定。结果经酶切鉴定和测序证实,人sTNFR1基因被正确克隆入真核表达载体pCDNA3.1(-)。结论真核表达质粒pCDNA3.1(-)-sTNFR1的构建为今后的研究打下了基础。

[Objective] To construct an eukaryotic expression vector of human sTNFR1 as a basis for further study. [Methods] The total RNA was extracted from Hela cells and used as a template to amplify human sTNFR1 gene by reverse transcription polymerase chain reaction （RT-PCR）. The PCR products were cloned into T vector and sub-cloned into vector pCDNA3.1（-）, an eukaryotic expression vector. The exactitude of recombinant vector pCD- NA3.1（-）-sTNFR1 was identified by digestion and DNA sequencing analysis. [Results] The recombinant vector pCDNA3.1 （-）-sTNFR1 was constructed successfully by DNA sequencing analysis and by digestion identification. [Conclusion] We constructed the eukaryotic expression vector containing Human sTNFR1 gene successfully, which is the foundation for further study.