CXCL16/SR-PSOX基因克隆与原核表达的研究  

Prokaryotic expression of human CXCL16/SR-PSOX fusion protein and its identification

作　　者：李丹[1] 陈兴明[1] 许丽艳[1] 易勇[1] 刘春雷[1] 肖敏[1] 敬华[1] 

机构地区：[1]解放军306医院检验科,北京100101

出　　处：《总装备部医学学报》2010年第4期191-194,共4页Medical Journal of General Equipment Headquarters

摘　　要：目的构建GST-CXCL16/SR-PSOX融合蛋白原核表达载体并进行原核表达及鉴定。方法从人外周血单个核细胞(PBMCs)中提取总mRNA,反转录得到cDNA后PCR扩增获得CX-CL16/SR-PSOX基因全长,构建融合表达载体pGST-CXCL16/SR-PSOX,通过酶切、测序及诱导表达后的免疫印迹实验验证重组载体构建的正确性及融合蛋白表达的正确性。结果 PCR扩增获得预期长度的DNA片段,克隆至pGEX-6p-1载体后测序与GeneBank相符。诱导表达获得的蛋白分子量与预期一致,免疫印迹实验证实为GST融合蛋白。结论 GST-CXCL16/SR-PSOX融合蛋白表达载体构建成功,原核表达顺利。Objective To construct a prokaryotic expression vector of GST-CXCL16/SR-PSOX. Methods The total mRNA was extracted from peripheral blood mononuclear cells,and reverse transcription polymerase chain reaction was performed to amplify the full length of CXCL16/SR-PSOX gene. The pGST-CXCL16/SR-PSOX was constructed by ligasing the CXCL16/SR-PSOX gene and pGEX-6p-1. The correctness of recombinant plasmid was verified by enzyme incision and sequencing,and the correctness of expression was verified by electrophoresis and Western blot. Results A specific fragment was amplified by PCR,and then cloned into pGEX-6p-1.The homology of sequence was confirmed by comparison to that in the GeneBank.The expression of GST-CXCL16/SR-PSOX fusion protein was confirmed by Western blot by using anti-GST antibody. Conclusion The prokaryotic expression vector pGST-CXCL16/SR-PSOX is successfully constructed,and the target fusion protein can be well expressed in BL21.

关 键 词：GST融合蛋白 CXCL16/SR-PSOX 原核表达 

分 类 号：R341[医药卫生—基础医学] Q78[生物学—分子生物学]