| 摘要: |
| 运用同源克隆方法, 从军曹鱼脑垂体中分离了生长激素基因(GH)的全长 cDNA。结果表明, 该基因 cDNA 全长 862bp, 含 615bp 的开放阅读框(ORF), 编码 204 个氨基酸; 其中包括 22 个氨基酸的信号肽和 182 个氨基酸的成熟肽。采用 T-A 克隆构建 pMD18-T-CoGH 重组质粒, 测序正确后经NdeⅠ和 XhoⅠ双酶切获得 CoGH 片段, 插入表达载体 pET-28a, 获得 pET-CoGH 重组质粒并在表达宿主菌
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| 关键词: 军曹鱼, 生长激素基因, cDNA 克隆, 原核表达 |
| DOI：10.11693/hyhz201103011011 |
| 分类号: |
| 基金项目:国家“十一五”科技支撑计划项目, 2007BAD29B03 号; 国家自然科学基金项目, 30972253 号; 广东省自然科学基金项目, 8452408801001272 号。 |
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| CLONING OF cDNA FOR RACHYCENTRON CANADUM GROWTH HORMONE AND ITS EXPRESSION IN PROKARYOCYTE |
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HAO Yu1,2, DENG Si-Ping1,2, LIU Li1,2, LIU Chu-Wu1,2
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1.Fisheries College, Guangdong Ocean University;2.Key Laboratory of Aquaculture in South China Sea for Aquatic Economic Animal of Guangdong Higher Education Institutes, Guangdong Ocean University
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| Abstract: |
| The full length cDNA encoding growth hormone of Rachycentron canadum was amplified and cloned from total RNA isolated from pituitary gland by RT-PCR. The sequence was 862 nucleotides long, including an ORF of 615 base pairs. The encoded protein contains 204 amino acids, comprising a 22 amino acids signal peptide and a 182 amino acids mature protein. The resulting cDNA sequence was undergone sequence analysis and subcloned into expression vector pET-28a between NdeⅠand XhoⅠrestriction sites and expressed in Escherichia coli BL21(DE3) cells. SDS-PAGE and Western-blotting analysis showed that the expressed protein accumulated as inclusion bodies and the molecular weight of expressed fusion protein was 22.4kDa. |
| Key words: Rachycentron canadum, Growth hormone gene, cDNA cloning, Prokaryotic expression |
