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中国农学通报

中国农学通报 ›› 2011, Vol. 27 ›› Issue (23): 6-11.

所属专题： 生物技术； 畜牧兽医

• 畜牧 动物医学 蚕 蜂 • 上一篇    下一篇

猪ATF4基因真核超表达载体的构建及鉴定

陈超 吴望军 熊远著   

  • 收稿日期:2011-01-19 修回日期:2011-03-03 出版日期:2011-09-15 发布日期:2011-09-15
  • 基金资助:

    国家重点基础研究发展规划(973 计划)

Construction of pcDNA3.1(+)-ATF4 of pig and identification

  • Received:2011-01-19 Revised:2011-03-03 Online:2011-09-15 Published:2011-09-15

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摘要：

摘 要：通过克隆猪ATF4基因CDS，构建pcDNA3.1(+)-ATF4真核超表达载体，为下一步在细胞水平和个体水平研究该基因功能奠定条件。提取RNA,运用RT-PCR技术和巢式PCR技术扩增猪ATF4全部编码序列，克隆转化到pMD18-T载体中，测序证实后，在克隆至真核超表达载体pcDNA3.1(+),构建了pcDNA3.1(+)-ATF4真核超表达载体，并进行酶切和测序鉴定。在细胞水平上进行转染鉴定。成功克隆了ATF4基因，并构建了pcDNA3.1(+)-ATF4真核超表达载体，瞬时转染C2C12细胞，超表达效果明显。为进一步研究该基因的功能奠定了基础。

关键词: 成长理论, 成长理论

Abstract:

Abstract: To construct an eukaryotic expression vector pcDNA3.1(+)-ATF4 by cloning mouse ATF4 gene and provide basis for establishment of gene function at the cellular and individual level.Extracted RNA, and using RT-PCR, and nested PCR method ,we successfully amplified coding sequence of porcine ATF4 and cloned it into the pMD18-T vector and sequencing confirmed and then constructed pcDNA3.1 (+)-ATF4 vector which was identified by restriction enzyme analysis and DNA sequencing.The pig ATF4 gene is successfully cloned and an eukaryotic expression vector pcDNA3.1-ATF4 is successfully constructed. Transiently transfected C2C12 cells and effect of expression is obvious, Which will contribute to further studies on the ATF4 function and to the establishment of condition.