【作者】 杨爱龙； 李芳秋； 张春华； 缪家文；

【Author】 YANG Ai-Long, LI Fang-Qiu, ZHANG Chun-Hua, MIAO Jia-Wen(Center of Medical Laboratory Sciences of PLA, Nanjing General Hospital of Nanjing Command, Nanjing 210002, China)

【机构】 南京军区南京总医院全军医学检验中心；

【摘要】 目的克隆人精子蛋白17(hSp17)基因,构建重组表达载体并表达重组蛋白。方法用RT-PCR法从人睾丸中克隆hSp17基因,构建重组表达质粒pET-28a(+)/hSp17,转化大肠杆菌BL21(DE3),IPTG诱导表达,SDS-PAGE检测,用特异性hSp17的mAb进行Western blot鉴定,Ni-NTA His-Bind树腊纯化重组蛋白。结果:克隆到hSp17 cDNA(456 bp)并经过DNA测序证实,表达和纯化得到重组蛋白hSp17(表观分子量为27 500),并经过Western blot鉴定。结论:成功克隆和表达hSp17基因。更多还原

【Abstract】 Objective To clone human sperm protein 17 (hSp17) cDNA and construct expression vector producing recombinant pro-tein.Methods hSp 17 cDNA amplified by RT-PCR from human testis was inserted into plasmid pET-28a(+). Plasmid pET-28a(+)/hSp17 transformed E.coli BL21 (DE3) . Recombinant protein hSp17 was produced under the induction of IPTG, analyzed by SDS-PAGE, identified by Western blot with specific mAb and purified by Ni-NTA His-Bind resin.Results The cDNA of hSp17 was cloned and se-quenced. Recombinant protein (apparent MT 27 500) was highly expressed and demonstrated by Western blot Conclusion: hSp17 gene was cloned and expressed successfully.更多还原