猪生长激素cDNA克隆及在大肠杆菌中的表达  

Cloning of cDNA for Porcine GH and Its Expression in E. coli

作　　者：白俊杰[1] 叶星[1] 李新辉[1] 简清[1] 罗建仁[1] 

机构地区：[1]中国水产科学研究院珠江水产研究所热带亚热带鱼类选育与养殖农业部重点开放实验室,广州510380

出　　处：《华中农业大学学报》2001年第2期99-102,共4页Journal of Huazhong Agricultural University

基　　金：农业部"九五"重点渔业科技项目 !(95 B 96 0 2 0 8 0 2 ) ;中国水产科学研究院重点基金! (99 0 8 0 2 )资助项目

摘　　要：用逆转录 聚合酶链式反应 (RT PCR)方法 ,从猪脑垂体总RNA中扩增出编码猪生长激素 (GH)成熟肽基因序列 ,定向克隆至质粒pUC18。序列分析表明 ,克隆的猪GHcDNA长 5 73bp ,不含信号肽序列 ,并在该序列之前加入一起始密码子ATG。将猪GHcDNA定向克隆至原核表达载体pBV2 2 0 ,构建成重组猪GH基因表达载体pBVpGH7。SDS PAGE和薄层扫描分析表明 :经 42℃诱导 ,pBVpGH7在大肠杆菌中可表达一分子量约 2 2 0 0 0的特异蛋白 ,表达量约占细胞总蛋白的 2 0 .5 %。Total RNA was isolated from porcine pituitary. The cDNA encoding mature growth hormone (GH) peptide was amplified by reverse transcription polymerase chain reaction (RT PCR) method using isolated total RNA as template. The amplified cDNA fragment was inserted into plasmid pUC18. The cloned cDNA has 573 bp and has no signal peptides sequence. A start codon ATG was added to the front of first amino acid code of mature GH sequence. The cDNA was subcloned into the downstream of the PR PL promoter of expression plasmid pBV220 and expression plasmid pBVpGH7 was constructed. SDS PAGE shows that E. coli DH5(containing pBVpGH7) has expressed a 22kD molecular weight specific protein when induced for 4 h at 42℃ and the proportion of expressed GH in total bacterial protein is about 20.5%.

关 键 词：猪 生长激素CDNA 基因克隆 大肠杆菌表达 鱼用饲料添加剂 促生长作用 RT-PCR 

分 类 号：S963.73[农业科学—水产养殖] Q78[农业科学—水产科学]