摘要
采用聚合酶链反应 (PCR)技术对鲤和鲑生长激素cDNA的 5’端和 3’端进行定向改造。将改造后的5种基因分别克隆到大肠杆菌表达质粒 pBV2 2 0进行原核表达 ,以研究鱼生长激素基因在大肠杆菌中的表达水平。实验结果表明 :(1)核糖核蛋白体结合位点 (SD)与起始密码子AUG之间的距离对鱼生长激素的表达水平有影响 ;(2 )鲑生长激素基因的终止密码子UAG可能不会有效终止该基因在大肠杆菌中翻译的进行而造成部分通读 ,加入大肠杆菌强终止码UAA可避免通读和产生超长蛋白 ;(3)对鲤和鲑生长激素cDNA的 5’端和 3’端进行相同的修饰 ,但两者生长激素的表达量却有很大差异 ,说明基因的密码子偏性对表达水平有影响 ;(4 )提高
In order to study the expression level of fish growth hormone (fGH) in Escherichia coli, common carp and salmon GH cDNA's 5'ends and 3'ends were modified by using polymerase chain reaction (PCR) methods. Five of modified fGH cDNAs were coloned to E.coli expression vector pBV220. The experiments showed that the distance between SD sequence and initiator AUG is concerned with fGH expression level; salmon GH cDNA's stop codon UAG may not stop E. coli's translation effectively, but the strong stop condon UAA of E. coli added can avoid expression of super long proteins; although carp GH and salmon GH cDNA's 5'and 3'ends are modified with the same sequences,the expression levels of the 2 fish GH are quite different, showing that the number and distribution of rare codon in fGH genes may affect expression level of fGH in E.coli; the free energy of RNA 5'ends and local codon adaptation index also affect gene expression.
出处
《中国水产科学》
CAS
CSCD
2000年第1期31-34,共4页
Journal of Fishery Sciences of China
基金
农业部"九五"重点渔业科技项目 (渔 95 -B - 96 - 0 2- 0 2 - 0 4 )
中国水产科学研究院基金!资助项目 (95 - 0 8- 0 1)
关键词
鱼生长激素
基因改造
表达水平
大肠杆菌
fish growth hormone
gene modification
expression level
Escherichia coli
RNA secondary structure
