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| 新城疫病毒Roakin株HN基因的克隆及真核表达载体的构建 | |
| 引用本文： | 张继乐,陈豪泰,丁耀忠,孙德惠,马丽娜,张杰,金雷,高云英,刘永生.新城疫病毒Roakin株HN基因的克隆及真核表达载体的构建[J].西北农业学报,2009,18(2):19-23,37. |
| 作者姓名： | 张继乐 陈豪泰 丁耀忠 孙德惠 马丽娜 张杰 金雷 高云英 刘永生 |
| 作者单位： | 1. 西北农林科技大学,动物医学院,陕西杨凌,712100;中国农业科学院,兰州兽医研究所,家畜疫病病原生物学国家重点实验室,农业部畜禽病毒学重点开放实验室,甘肃兰州,730046 2. 中国农业科学院,兰州兽医研究所,家畜疫病病原生物学国家重点实验室,农业部畜禽病毒学重点开放实验室,甘肃兰州,730046 3. 西北农林科技大学,动物医学院,陕西杨凌,712100 |
| 基金项目： | 中国农业科学院兰州兽医研究所所长基金 |
| 摘 要： | 克隆新城疫病毒(Newcastle disease virus,NDV)Roakin株HN基因,并构建真核表达载体.应用RT-PCR扩增出NDV Roakin株HN基因,将其克隆入pMD18-T载体,并进行了测序鉴定和HN基因的序列分析.然后分别将NDV的HN基因和选择标记基因-二氢叶酸还原酶(dhfr)基因插入到真核表达载体pCI-neo中,通过PCR扩增、酶切分析和测序分析等鉴定所构建的重组真核表达载体.RT-PCR扩增的NDV Roakin株HN序列与GenBank中标准毒株(登录号AY289000)HN序列同源性为99.8%,构建了HN基因的真核表达质粒pCI-HN-D.成功构建了含有NDV Roakin株HN基因和dhfr基因的真核表达载体. |
| 关 键 词： | 新城疫病毒 HN基因 dhfr基因 真核表达载体 |
| 收稿时间： | 2008/9/23 0:00:00 |
| 修稿时间： | 2008/12/1 0:00:00 |
Cloning and Construction of Eukaryotic Expression Vector of HN Gene of Newcastle Disease Virus Roakin Strain |
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| ZHANG Jile,CHEN Haotai,DING Yaozhong,SUN Dehui,MA Lin,ZHANG Jie,JIN Lei,GAO Yunying and LIU Yongsheng.Cloning and Construction of Eukaryotic Expression Vector of HN Gene of Newcastle Disease Virus Roakin Strain[J].Acta Agriculturae Boreali-occidentalis Sinica,2009,18(2):19-23,37. | |
| Authors: | ZHANG Jile CHEN Haotai DING Yaozhong SUN Dehui MA Lin ZHANG Jie JIN Lei GAO Yunying and LIU Yongsheng |
| Institution: | College of Veterinary Medicine, Northwest A&F University, Yangling Shaanxi 712100, China;Key Laboratory of Animal Virology of, Ministry of Agriculture/State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou Gansu 730046, China;Key Laboratory of Animal Virology of, Ministry of Agriculture/State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou Gansu 730046, China;Key Laboratory of Animal Virology of, Ministry of Agriculture/State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou Gansu 730046, China;Key Laboratory of Animal Virology of, Ministry of Agriculture/State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou Gansu 730046, China;Key Laboratory of Animal Virology of, Ministry of Agriculture/State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou Gansu 730046, China;Key Laboratory of Animal Virology of, Ministry of Agriculture/State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou Gansu 730046, China;Key Laboratory of Animal Virology of, Ministry of Agriculture/State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou Gansu 730046, China;College of Veterinary Medicine, Northwest A&F University, Yangling Shaanxi 712100, China;Key Laboratory of Animal Virology of, Ministry of Agriculture/State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou Gansu 730046, China |
| Abstract: | To clone The HN gene of Newcastle disease virus (NDV) Roakin strain and construct a eukaryotic expression vector which can be expressed in Chinese Hamster Ovary Cells CHO/dhfr-.The HN gene of Newcastle disease virus (NDV) Roakin strain was amplified by RT-PCR and cloned into pMD18-T vector and sequenced.The HN gene of the virus and dhfr gene was cloned into high efficient eukaryotic vector pCt-neo.PCR amplification, restriction emzyme digestion and sequencing analysis was applied to identify the recombinant expression plasmids.The sequence homology of HN cloned with the HN(AY289000)on GenBank was 99.8%;Mammalian recombinant expression plasmid pCI-HN-D was constructed.Recombinant expression plasmid pCI-HN-D harboring dhfr gene and HN gene of NDV has been successfully constructed and lay the foundation of high-level expression of HN urotein of NDV in mammalian cell CHO/dhfr-. |
| Keywords: | NDV HN gene dhfr gene Eukaryotic expression vector |
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