中文摘要：

目的原核表达人增生性瘢痕新候选相关蛋白P311，并对该重组蛋白进行纯化、鉴定。方法通过PCR方法扩增人P311基因，利用BamHⅠ和XhoⅠ酶切位点将其克隆至谷胱甘肽-S-转移酶（glutathione-s-transferase，GST）融合蛋白表达载体pGEX-4T-1中，转化大肠杆菌BL21（DE3）后，经异丙基硫代-β-D-半乳糖苷（isopropyl-β-D-5-thiogalactoside，IPTG）诱导表达，用SDS-PAGE、Western blotting等方法鉴定表达产物，并用谷胱甘肽-琼脂糖小珠亲和纯化表达的GST-P311蛋白。结果重组载体pGEX-4T-P311经酶切与测序鉴定证实构建成功。导人大肠杆菌进行表达，表达产物相对分子量为34KD左右，与预期值相符。该条带经免疫印迹检测鉴定为GST抗体阳性，获得了纯化的GST-P311蛋白。结论构建了pGEX-4T-P311原核表达载体，并在大肠杆菌BL21（DE3）中成功表达，亲和纯化获得较高纯度的GST融合蛋白，为下一步继续研究P311的生物学功能奠定了基础。

英文摘要：

Objective To express, purify and identify human P311 protein, a candidate protein in hypertrophic scar formation. Methods The cDNA sequence of human P311 obtained by PCR was cloned into the prokaryotic expression vector pGEX-4T-1 containing glutathione-s-transferase （GST）. pGEX-4T-P311 was transformed into E. coli BL21 （DE3）. The expression of GST-P311 fusion protein was induced by IPTG, identified by both SDS-PAGE and Western blotting, and then purified with glutathione-sepharose beads. Results The restriction endonuclease digestion and the sequencing of recombinant plasmid demonstrated that pGEX-4T- P311 vector was successfully constructed. The relative molecular weight of expressed protein was about 34×10^3 which was as the same as the objective protein, and the corresponding protein was identified by the immunoblotting with antibody against GST. Conclusion pGEX-4T-P311 vector is correctly constructed, and the GST-P311 fusion protein is successfully expressed in E. coli BL21 （ DE3 ） and purified.