山羊PTHrP基因CDs区克隆及其融合蛋白的表达  

Cloning of Capra hircus PTHrP and its prokaryotic expression in Escherichia coli

作　　者：杨振宇[1] 郑惠玲[1] 邢瑞芳[1] 祝珍珍[1] 安俊辉[1] 

机构地区：[1]西北农林科技大学动物科技学院,陕西省农业分子生物学重点实验室,陕西杨凌712100

出　　处：《西北农林科技大学学报（自然科学版）》2010年第10期45-50,共6页Journal of Northwest A&F University(Natural Science Edition)

基　　金：西北农林科技大学科研启动基金项目(08080112);农业部公益性行业科研专项(3-45)

摘　　要：【目的】克隆山羊甲状旁腺相关肽(Parathyroid hormone related protein,PTHrP)基因,并进行原核表达,获得具有生物学活性的融合蛋白。【方法】无菌采集山羊乳腺组织,采用TRIZOL法提取总RNA,RT-PCR克隆山羊PTHrP基因全CDs区。将山羊PTHrP基因全CDs区插入pET-32a(+)表达质粒构建pET-PTHrP质粒,经酶切和测序鉴定后,将其转化E.coliBL21(DE3)菌株,经2mmol/LIPTG、37℃诱导表达8h后,进行SDS-PAGE和Westernblotting分析。【结果】获得了山羊PTHrP基因全CDs区,长534bp(GenBank登录号GU573787);酶切和测序显示,原核表达载体pET-PTHrP构建成功;SDS-PAGE和Westernblotting分析表明,在E.coliBL21(DE3)中成功诱导表达了pET-PTHrP融合蛋白。【结论】克隆了山羊PTHrP基因,获得了PTHrP融合蛋白。【Objective】This experiment was conducted to study Capra hircus parathyroid hormone related protein PTHrPgene expression in E.coli under the condition of cloning PTHrPgene complete CDs sequence.【Method】Total RNA was isolated fromC.hircus mammary gland tissues.The CDs sequnence of PTHrPgene were amplified by RT-PCR.CDs of PTHrPwas inserted to the prokaryotic expression vector pET-32a(+) and expressed(2mmol/L IPTG for 8hat 37℃)in E.coli BL21(DE3)host cells.The results were analysed by SDS-PAGE and Western bolt...

关 键 词：PTHrP基因 原核表达 蛋白纯化 山羊 

分 类 号：S827[农业科学—畜牧学]