广西巴马小型猪ANK1基因启动子克隆及其活性分析

发布时间：2018-04-25 09:39

  本文选题：广西巴马小型猪 + 锚蛋白(ANK)　； 参考：《南方农业学报》2017年04期

【摘要】：【目的】克隆广西巴马小型猪ANK1基因启动子,确定其活性核心区,为研究ANK1基因启动子与肉质性状的相关性及构建动物疾病模型打下基础。【方法】通过在线软件对ANK1基因启动子的转录因子结合位点进行预测,根据转录因子结合位点设计特异引物扩增不同长度的ANK1基因启动子片段,并利用双荧光素酶试剂盒检测其荧光值,以确定不同ANK1基因启动子片段的活性。【结果】发现ANK1基因启动子存在1个转录起始位点(TSS)、2个Cp G岛和多个转录因子结合位点,并以此作为ANK1基因启动子的分段依据,将其分割为P638、P791、P1113、P1163、P1648、P1694、P1796和P2074等8个不同长度的目的片段。成功克隆获得的8个ANK1基因启动子片段经KpnⅠ和HindⅢ双酶切、T4真核表达载体连接、细胞转染等方法构建8个双荧光素酶重组报告基因,双荧光素酶试剂盒检测结果显示,广西巴马小型猪ANK1基因启动子在P1796片段活性最强,与其他片段存在显著差异(P0.05)。【结论】成功克隆获得广西巴马小型猪ANK1基因启动子的8个片段,且利用双荧光素酶试剂盒检测确定其核心启动子区域出现在P1796片段。
[Abstract]:[objective] to clone the promoter of ANK1 gene from Guangxi Bama miniature pig and determine its active core region. In order to study the correlation between ANK1 gene promoter and fleshy traits and to construct animal disease model, the transcription factor binding sites of ANK1 gene promoter were predicted by online software. According to the transcription factor binding site, specific primers were designed to amplify the promoter fragments of ANK1 gene of different lengths, and the fluorescence values of the promoter fragments were detected by double luciferase kit. In order to determine the activity of different promoter fragments of ANK1 gene. [results] it was found that there were one transcriptional initiation site (TSS), two CP G islands and multiple transcription factor binding sites in the promoter of ANK1 gene, which were used as the segmental basis of the promoter of ANK1 gene. It was divided into P638P791P1113P1163, P1648P1694, P1796 and P2074, etc. Eight ANK1 promoter fragments were successfully cloned and ligated into Kpn 鈪，

本文编号：1800804