TNF受体(P55)胞外区和IgGFc嵌合蛋白高效原核表达载体的构建  

Construction of high efficiency prokaryotic expression vector of extra-cellular domain of TNFR and IgGFc chimeric protein

作　　者：祖东[1] 徐春晓[2] 张智清[2] 谭锦泉[1] 

机构地区：[1]安徽医科大学免疫学教研室,合肥230032 [2]中国预防医学科学院病毒学研究所基因室,北京100052

出　　处：《安徽医科大学学报》2002年第2期89-92,共4页Acta Universitatis Medicinalis Anhui

基　　金：国家 8 6 3生物领域基金资助项目 (编号 :2 0 0 1AA2 15 2 5 1)

摘　　要：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。Objective To construct high efficiency prokaryotic expression vector of extra-cellular domain of TNFR and IgGFc chimeric protein. Methods Complete extra-cellular domain of TNFR gene was amplified from human heart cDNA library by PCR, and then cloned into pGEM-T/IgGFc vector. With this template, no signal peptide of extra-cellular domain of TNFR and IgGFc chimeric gene was amplified with high efficiency prokaryotic codons in the upstream primer, then it was subcloned into pUC19 vector. After sequencing, TNFR∶IgGFc chimeric gene was cloned into the prokaryotic expression vector pBV220. Results Prokaryotic high efficiency expression vector of extra-cellular domain of TNFR and IgGFc chimeric protein was obtained successfully, named as pBV220/TNFR-ED∶IgGFc. Conclusion Construction of pBV220/TNFR-ED∶IgGFc will provide the basis for expression of TNFR-ED∶IgGFc chimeric protein and for further studying its function in the neutralization the side effects of TNF or treatment of autoimmune diseases.

关 键 词：TNF受体 IgGFc 嵌合蛋白 高效原核表达 载体 构建 

分 类 号：R392.11[医药卫生—免疫学] Q786[医药卫生—基础医学]