Prokaryotic expression, purification and identification of human P311 protein

谭江琳; 袁顺宗; 彭旭; 马兵; 罗高兴; 吴军

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Prokaryotic expression, purification and identification of human P311 protein

更新时间：2026-01-09

- Prokaryotic expression, purification and identification of human P311 protein
- Journal of Army Medical University Vol. 30, Issue 1, Pages: 19-22(2008)
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  CLC： R341;R394-33
- Published：2008
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谭江琳,袁顺宗,彭旭,马兵,罗高兴,吴军.人重组蛋白P311的原核表达纯化与鉴定[J].陆军军医大学学报(原第三军医大学学报),2008,30(01):19-22.

　TAN Jiang-lin,YUAN Shun-zong,PENG Xu,MA Bing,LUO Gao-xing,WU Jun.Prokaryotic expression, purification and identification of human P311 protein[J].J Amry Med Univ (J Third Mil Med Univ),2008,30(01):19-22.
谭江琳,袁顺宗,彭旭,马兵,罗高兴,吴军.人重组蛋白P311的原核表达纯化与鉴定[J].陆军军医大学学报(原第三军医大学学报),2008,30(01):19-22. DOI：

　TAN Jiang-lin,YUAN Shun-zong,PENG Xu,MA Bing,LUO Gao-xing,WU Jun.Prokaryotic expression, purification and identification of human P311 protein[J].J Amry Med Univ (J Third Mil Med Univ),2008,30(01):19-22. DOI：

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。导入大肠杆菌进行表达，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。

Abstract

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

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.

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