论文部分内容阅读
目的 构建TNF受体 (P5 5 )胞外区和IgGFc嵌合蛋白的高效原核表达载体。方法 以人心肌cDNA文库为模板 ,PCR扩增TNF受体 (P5 5 )胞外区全基因 (sTNFR ED) ,亚克隆入pGEM T/IgGFc中构建 pGEM T/sTNFR ED∶Ig GFc重组子 ,然后以其为模板 ,在上游引物中引入原核细胞高频密码子 ,再次PCR扩增去信号肽TNF受体 (P5 5 )胞外区和IgGFc嵌合基因 (TNFR ED∶IgGFc) ,亚克隆入 pUC19载体 ,经序列测定正确后克隆入原核表达载体 pBV2 2 0中。结果 成功构建了TNF受体 (P5 5 )胞外区和IgGFc嵌合蛋白高效原核表达载体 ,命名为 pBV2 2 0 /TNFR ED∶IgGFc。结论 pBV2 2 0 /TNFR ED∶IgGFc表达载体的构建 ,为进一步表达TNF受体 (P5 5 )胞外区和IgGFc嵌合蛋白及其在中和TNF毒副作用、治疗自身免疫性疾病中的作用机制研究奠定了基础
Objective To construct a prokaryotic expression vector for the extracellular domain of TNF receptor (P5 5) and IgGFc chimeric protein. Methods Human cardiac cDNA library was used as a template to amplify sTNFR ED of TNF receptor (P5 5) gene by PCR and subcloned into pGEM T / IgGFc to construct pGEM T / sTNFR ED: IgG GF recombinants. Using it as a template, high-frequency codons of prokaryotic cells were introduced into the upstream primer, and the extracellular region of the signal peptide TNF receptor (P5 5) and the IgGFc chimeric gene (TNFR ED: IgGGF) were again PCR-amplified and subcloned into pUC19 The vector was sequenced and cloned into prokaryotic expression vector pBV220. Results The extracellular domain of TNF receptor (P5 5) and the prokaryotic expression vector of IgGFc chimeric protein were successfully constructed and named as pBV2 2 0 / TNFR ED: IgGFc. Conclusion The construction of pBV2 2 0 / TNFR ED: IgGFc expression vector is to further express the extracellular domain of TNF receptor (P5 5) and the IgGFc chimeric protein and its mechanism of action in neutralizing TNF toxicity and treating autoimmune diseases Research laid the foundation