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| 猪GM-CSF基因的克隆及其真核表达载体的构建 | |
| 引用本文： | 黄海军,高其双,钱运国,邵淑敏,黄远全,陈志华,陶弼菲,向敏,占才耀,夏瑜,王连芳,任远.猪GM-CSF基因的克隆及其真核表达载体的构建[J].生物技术通报,2011(4). |
| 作者姓名： | 黄海军 高其双 钱运国 邵淑敏 黄远全 陈志华 陶弼菲 向敏 占才耀 夏瑜 王连芳 任远 |
| 作者单位： | 1. 武汉市畜牧兽医科学研究所,武汉,430065 2. 华中农业大学农业部猪遗传育种重点开放实验室农业动物遗传育种与繁殖教育部重点实验室,武汉,430070 3. 湖北省赤壁市动物疫病预防控制中心,成宁,437300 |
| 基金项目： | 武汉市农科院创新项目，国家"863"课题项目 |
| 摘 要： | 采用PCR方法从猪外周血液淋巴细胞cDNA中扩增出与预期设计大小相符的GM-CSF基因特异性条带,PCR产物经EcoR Ⅰ和Xho Ⅰ双酶切后,插入到载体pIRES2-EGFP构建成真核表达载体pIRES2-EGFP-GM-CSF.经PCR鉴定、限制性内切酶酶切分析和克隆片段序列测定、比较,证实了重组质粒的正确性.将构建好的真核表达质粒转染到山羊胎儿成纤维细胞中进行瞬时表达,荧光检测证实细胞转染成功.pIRES2-EGFP-pGM-CSF真核表达载体的成功构建为下一步在细胞水平研究GM-CSF蛋白功能以及进一步将其开发为高档疫苗佐方剂奠定了基础. |
| 关 键 词： | 猪粒细胞-巨噬细胞集落刺激因子 克隆 真核表达载体 瞬时转染 |
Cloning and Construction of Eukaryotic Expression Vector for GM-CSF of Porcine |
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| Huang Haijun,Gao Qishuang,Qian Yunguo,Shao Shumin,Huang Yuanquan,Chen Zhihua,Tao Bifei,Xiang Min,Zhan Caiyao,Xia Yu,Wang Lianfang,Ren Yuan.Cloning and Construction of Eukaryotic Expression Vector for GM-CSF of Porcine[J].Biotechnology Bulletin,2011(4). | |
| Authors: | Huang Haijun Gao Qishuang Qian Yunguo Shao Shumin Huang Yuanquan Chen Zhihua Tao Bifei Xiang Min Zhan Caiyao Xia Yu Wang Lianfang Ren Yuan |
| Institution: | Huang Haijun1 Gao Qishuang1 Qian Yunguo1 Shao Shumin 2 Huang Yuanquan3 Chen Zhihua1 Tao Bifei1 Xiang Min1 Zhan Caiyao1 Xia Yu1 Wang Lianfang1 Ren Yuan1(1 Wuhan Institute of Animal and Veterinary Science,Wuhan 430065,2 Key Laboratory of Swine Breeding and Genetics,Ministry of Agriculture & Key Laboratory of Agricultural Animal Genetics,Breeding and Reproduction,Huazhong Agricultural University,Ministry of Education,Wuhan 430070,3 Chibi Center for Control and Prevention of Aquatic Animal Infectious Disease,Xi... |
| Abstract: | The whole cDNA sequence encoding porcine granulocyte-macrophase colony-stimulating factor(pGM-CSF)was amplified by PCR from peripheral blood lymphocytes of porcine.The PCR product was digested with EcoR I plus Xho I,then inserted into pIRES2-EGFP between the Xho I and EcoR I sites to generate a eukaryotic expression plasmid pIRES2-EGFP-pGM-CSF.The recombinant colonies were identified by restriction enzyme digestion,PCR and sequencing.These results showed that,the eukaryotic expression plasmid pIRES2-EGFP-pG... |
| Keywords: | Porcine granulocyte-macrophage colony-stimulating factor Cloning Eukaryotic expression vector Transient transfection |
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