人血浆蛋白S成熟肽基因在大肠杆菌中的融合表达  

GST fusion expression of the mature peptide of human protein S in E.coli

作　　者：卢爱薇[1] 兰风华[1] 程烽[1] 郑德柱[1] 谢飞[1] 朱忠勇[1] 

机构地区：[1]南京军区福州总医院全军医学检验中心,350025

出　　处：《临床输血与检验》2004年第1期12-15,共4页Journal of Clinical Transfusion and Laboratory Medicine

摘　　要：目的　构建人血浆蛋白 S的原核表达载体并诱导其表达。方法　自行设计引物 ,采用 PCR法 ,以现有人血浆蛋白 S真核表达载体为模板 ,扩增人血浆蛋白 S成熟肽的编码序列。 PCR产物经 Eco RI和 Bam HI双酶切后 ,克隆至 GST融合表达载体 p GEX-2 T中 ,在 E.coli BL2 1 中诱导 GST-人血浆蛋白 S融合蛋白的表达。结果　对重组质粒的序列分析表明 ,插入片段的序列与 Gen Bank登录的人血浆蛋白 S基因编码序列完全一致。 1 0 % SDS聚丙烯酰胺凝胶电泳显示 ,在IPTG的诱导下 ,BL2 1 重组菌高效表达分子量约为 96k D的产物 ,并可通过 GST亲和层析柱纯化。结论　人血浆蛋白 S编码序列已被克隆至 GST融合表达载体 p GEX-2 T,并在 E.coli BL2 1Objective To construct a prokaryotic vector for the expression of human protein S. Methods The coding sequence of protein S mature peptide was amplified from an existing recombinant plasmid (PS pcDNA3) with primers designed in our laboratory, and cloned into pGEX-2T after restrictive digestion with EcoRI and BamHI . GST-protein S fusion protein was expressed after induction by IPTG. Results Sequencing and restriction digestion of the recombinant plasmid revealed the existence of the coding sequence for protein S mature peptide. SDS-PAGE showed that a protein band of about 96 kD could be induced by IPTG in the recombinant bacteria harboring the plasmid. Conclusion The coding sequence of protein S mature peptide is introduced into the pGEX-2T plasmid and a GST-fused protein S mature peptide could be induced at a high level.

关 键 词：大肠杆菌 融合表达 分子克隆 蛋白S 

分 类 号：R392.11[医药卫生—免疫学]