| 牛eIF5基因原核表达与蛋白纯化 |
| |
| 引用本文： | 高学军,臧艳丽,魏诚杰,骆超超.牛eIF5基因原核表达与蛋白纯化[J].东北农业大学学报,2016(6):68-73. |
| |
| 作者姓名： | 高学军 臧艳丽 魏诚杰 骆超超 |
| |
| 作者单位： | 东北农业大学黑龙江省高校农业生物功能基因重点实验室,哈尔滨,150030 |
| |
| 基金项目： | 国家高技术研究发展计划(863)项目(2013AA102504-03) |
| |
| 摘 要： | 研究应用PCR方法扩增牛e IF5基因编码序列,通过Eco RⅠ和SalⅠ酶切位点将其定向插入p GEX-4T-1载体,构建原核表达重组质粒p GEX-4T-1-e IF5,并转化大肠埃希菌BL21。酶切和测序鉴定正确后,用异丙基硫代-β-D-半乳糖苷(IPTG)大量诱导表达,利用GST-Sepharose 4B亲和珠纯化,SDS-PAGE和Western Blot鉴定。结果表明,原核表达载体p GEX-4T-1-e IF5构建成功,Western Blot证实融合蛋白大量表达,纯化后获得GSTe IF5融合蛋白,为深入研究牛e IF5蛋白功能奠定基础。
|
| 关 键 词： | 牛 eIF5 克隆 原核表达 纯化 |
Prokaryotic expression and protein purification of gene bovine eIF5 |
| |
| Abstract: | The bovine eIF5 gene coding sequence was amplified by polymerase chain reaction (PCR) and was cloned into Eco R I and SalⅠ sites of pGEX-4T-1 vector to construct the recombinant plasmid pGEX-4T-1-eIF5. The recombinant plasmid was identified by restriction enzyme digestion and sequencing. Then it was transformed into BL21, induced by IPTG and purified by GST-Sepharose 4B beads, identified by SDS-PAGE and Western Blot analysis. Results showed that the prokaryotic expression vector pGEX-4T-1-eIF5 was successful y constructed. Western Blot confirmed the recombinant protein GST-eIF5 were expressed and purified. This study provided the basis for the further research on the protein function of bovine eIF5. |
| |
| Keywords: | bovine eIF5 cloning prokaryotic expression purification |
| 本文献已被 CNKI 万方数据 等数据库收录！ |
|
