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皮蝇Hypodermin A基因的克隆及表达质粒构建

Cloning and Construction of Recombinant Expression of Hypodermin A Tang

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【作者】 唐立刚杨东英罗建勋欧阳五庆殷宏

【Author】 Li-gang 1,2 Yang Dong-ying 1,2 Luo Jian-xun 2 Ouyang Wu-qing 1 Yin Hong 2 (1.College of Animal science and Technology,Northwest Sci-Tech University of Agriculture and Forestry,ShanXi Yangling712100;2.Lanzhou Veterinary Research Institute,Chinese Academy of Agricultural Sciences,GanSu Lanzhou,730046)

【机构】 西北农林科技大学动物科技学院中国农业科学院兰州兽医研究所中国农业科学院兰州兽医研究所 陕西杨陵712100中国农业科学院兰州兽医研究所甘肃兰州730046陕西杨陵712100中国农业科学院兰州兽医研究所甘肃兰州730046甘肃兰州730046陕西杨陵712100甘肃兰州730046

【摘要】 参照牛皮蝇HypoderminA(HA)基因的核苷酸序列,设计一对引物,以皮蝇总RNA为模板进行RT--PCR扩增牛皮蝇HA基因,将扩增基因进行克隆测序。序列分析表明,所克隆获得的基因与GenBank中已经登录的核苷酸和氨基酸的同源性分别为97.2%和99.3%。同时,将该基因与表达载体PGEX-4T-1连接,构建并获得阳性重组表达载体。

【Abstract】 Total RNA was extracted from Hypoderma,then mRNA was isolated by using oligo(dT)as a probe.The HA gene specific primers were devised by GOLDkey software.A700bp specific fragment was amplified by RT-PCR and ligated into PGEM-T Easy vector.It was identified by restriction endonuclease analysis and PCR and sequencing that this fragment contained the complete open reading frame (ORF)of the HA gene.In comparison with GenBank data,the homologies of the nucleotide sequence and amino acid sequence are97.2%and99.3%,respectively.After the HA-T was restrictively digested,the interested gene HA was subcoloned into the prokaryotic expression vector PGEM-4T-1.Positive clones were selected by restriction endonuclease analysis and PCR identification.

【基金】 国家863项目(2001AA249081);欧盟资助项目(ICA4-CT-2000-30028)
  • 【文献出处】 动物科学与动物医学 ,Animal Science and Veterinary Medicine , 编辑部邮箱 ,2004年01期
  • 【分类号】S852.7
  • 【被引频次】6
  • 【下载频次】74
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