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当前位置: 首页 > 文章 > 水牛抑制素α亚基基因的克隆与原核表达 黑龙江畜牧兽医 2009 (8) 14-17
Position: Home > Articles > Cloning and prokaryotic expression of swamp buffalo Inhibin alpha subunit gene Heilongjiang Animal Science and Veterinary Medicine 2009 (8) 14-17

水牛抑制素α亚基基因的克隆与原核表达

作  者：
谢体三;农微;张新民;黄雅琼;崔奎青;石德顺
单  位：
广西大学动物繁殖研究所
关键词：
抑制素α亚基;水牛;原核表达
摘  要：
采用RT-PCR方法从水牛卵巢总RNA中扩增抑制素α亚基基因,并克隆入pMD18-T载体,进行PCR、双酶切及测序鉴定。序列分析结果表明:水牛抑制素α亚基基因编码序列长为1083 bp,编码360个氨基酸,与牛、人、猪抑制素α亚基基因CDS成熟蛋白氨基酸的同源性分别为96%、80%、87%,表明抑制素α亚基是一组在进化上高度保守的蛋白质。将水牛抑制素α亚基基因CDS克隆到pET-30a表达载体中,转化宿主菌BL21(DE3)进行原核表达。在1 mmol/L IPTG中,37℃诱导表达4 h后抑制素α亚基基因重组蛋白可成功获得表达。将表达产物进行SDS-PAGE分析,结果表达产物主要以包涵体形式存在,分子质量约为40 ku。
译  名：
Cloning and prokaryotic expression of swamp buffalo Inhibin alpha subunit gene
作  者：
XIE Ti-san,NONG Wei,SHI De-shun,et al (Animal Reproduction Institute,Guangxi University,Nanning 530005,China)
关键词：
Inhibin alpha subunit;swamp buffalo;prokaryotic expression
摘  要：
The Inhibin alpha subunit gene was amplified by RT-PCR from total RNA of swamp buffalo ovary and cloned into pMD 18-T vector.The recombinant vector pMD 18-T-INHA was identified by PCR,restriction endonuclease analysis and sequenced.The INHA CDS was amplified and cloned into pET-30a(+) expression vector.The recombinant pET-30a-INHA expression plasmid was confirmed to be constructed successfully by restriction endonuclease analysis and nucleotide sequenced.Then the recombinant plasmid was transformed into BL21(DE3) and induced expression by IPTG.The result show the buffalo INHA CDS contain 1 083 bp,encoded 360 aa,and exhibited a high homology with bovine(96%),human(80%),pig(87%).This result indicated the INHA gene was relatively conservative in the progress of evolution.The recombinant protein INHA was expressed successfully when induced by 1 mmol/L IPTG at 37 ℃ for 4 hours.Most of the expression products were inclusion body with the molecular weigh of about 40 000 identified by SDS-PAGE.

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