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RTPCR法从人睾丸中克隆hSp17基因,构建重组表达质粒pET28a(+)/hSp17,转化大肠杆菌BL21(DE3),IPTG诱导表达,SDSPAGE检测,用特异性hSp17的mAb进行Westernblot鉴定,NiNTAHisBind树脂纯化重组蛋白.结果:克隆到hSp17cDNA(456bp)并经过DNA测序证实,表达和纯化得到重组蛋白hSp17(表观Mr为27500),并经过Westernblot鉴定.结论:成功克隆和表达hSp17基因.
AIM： To clone human sperm protein 17 （hSp17） cDNA and to construct expression vector producing recombinant protein. METHODS： hSp17 cDNA amplified by RT-PCR from human testis was inserted into plasmid pET- 28a（ ＋ ） and plasmid pET-28a （ ＋ ）/hSp17 transformed E. coli BL21 （DE3）. Recombinant protein hSpl？ 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 sequenced. Recombinant protein （apparent Mr 26 500） was highly expressed and demonstrated by Western blot. CONCLUSION： hSp17 gene has been successfully cloned and expressed.
出处
《第四军医大学学报》
北大核心
2005年第20期1892-1894,共3页
Journal of the Fourth Military Medical University
关键词
人精子蛋白17
克隆
分子
基因表达
印迹法
蛋白质
human sperm protein 17： cloning, molecular
gene expession
blotting
western
