广西巴马小型猪ANK1基因启动子克隆及其活性分析
【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...
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| Published in | 南方农业学报 Vol. 48; no. 4; pp. 716 - 720 |
|---|---|
| Main Author | |
| Format | Journal Article |
| Language | Chinese |
| Published |
广西大学 动物科学技术学院,南宁,530004
2017
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| Subjects | |
| Online Access | Get full text |
| ISSN | 2095-1191 |
| DOI | 10.3969/j.issn.2095-1191.2017.04.025 |
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| Abstract | 【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（P〈0.05）。【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。 |
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| AbstractList | S828.89; [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CpG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(P<0.05).[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. 【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（P〈0.05）。【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_FL | [Objective]In this study, ANK1 gene promoter of Guangxi Bama miniature pig was cloned, and its core ac-tivity areas were detected, so as to lay a foundation for researching correlation between ANK1 gene promoter and meat qual-ity traits, and establishing animal disease model. [Method]Transcription factor binding sites of ANK1 promoter were pre-dicted with online software, ANK1 promoter fragments of different lengths were amplified by specific primers based on tran-scription factor binding sites. To determine the activity of different fragments, fluorescence values of different promoter re-gions were tested using dual luciferase kits. [Result]One transcription starting sites (TSS), two CpG islands and multiple transcription factor binding sites were found in ANK1 promoter. The results served as basis for segmentation of ANK1 gene promoter and it was devided into eight target fragments of different length, namely P638,P791,P1113,P1163,P1648, P1694,P1796 and P2074. The eight segments was cloned. Through KpnⅠand HindⅢdouble enzyme digestion, T4 eukaryotic expression vector linkage and cell transfection, eight dual luciferase kit recombination reporter genes were established. Based on dual luciferase detection, the activity of ANK1 gene promoter was the strongest in P1796 fragment, and the difference with others fragments was significant (P<0.05). [Conclusion]In this study, eight fragments of the ANK1 gene promoter from Guangxi Bama miniature pig are successfully cloned and the core region of the ANK1 gene promoter is identified as P1796 fragment by dual luciferase kits. |
| Author | 李龙 司景磊 夏攀洁 綦文晶 龙开旭 夏利 何剑雄 吴敏 兰干球 |
| AuthorAffiliation | 广西大学动物科学技术学院,南宁530004 |
| AuthorAffiliation_xml | – name: 广西大学 动物科学技术学院,南宁,530004 |
| Author_FL | XIA Li WU Min LAN Gan-qiu SI Jing-lei QI Wen-jing XIA Pan-jie HE Jian-xiong LI Long LONG Kai-xu |
| Author_FL_xml | – sequence: 1 fullname: LI Long – sequence: 2 fullname: SI Jing-lei – sequence: 3 fullname: XIA Pan-jie – sequence: 4 fullname: QI Wen-jing – sequence: 5 fullname: LONG Kai-xu – sequence: 6 fullname: XIA Li – sequence: 7 fullname: HE Jian-xiong – sequence: 8 fullname: WU Min – sequence: 9 fullname: LAN Gan-qiu |
| Author_xml | – sequence: 1 fullname: 李龙 司景磊 夏攀洁 綦文晶 龙开旭 夏利 何剑雄 吴敏 兰干球 |
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| Copyright | Copyright © Wanfang Data Co. Ltd. All Rights Reserved. |
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| DocumentTitleAlternate | ANK1 gene promoter in Guangxi Bama miniature pig： Cloning and activity analysis |
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| Keywords | Guangxi Bama miniature pig 启动子 ankyrins(ANK) 广西巴马小型猪 ANK1基因 promoter 活性分析 activity analysis 锚蛋白(ANK) ANK1 gene |
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| Notes | Guangxi Bama miniature pig; ankyrins（ANK）; ANK1 gene; promoter; activity analysis 45-1381/S LI Long, SI Jing-lei, XIA Pan-jie, QI Wen-jing, LONG Kai-xu, XIA Li, HE Jian-xiong, WU Min, LAN Gan-qiu（College of Animal Science and Technology, Guangxi University, Nanning 530004, China） Objective]In this study, ANK1 gene promoter of Guangxi Bama miniature pig was cloned, and its core activity areas were detected, so as to lay a foundation for researching correlation between ANK1 gene promoter and meat quality traits, and establishing animal disease model. [Method]Transcription factor binding sites of ANK1 promoter were predicted with online software, ANK1 promoter fragments of different lengths were amplified by specific primers based on tran- scription factor binding sites. To determine the activity of different fragments, fluorescence values of different promoter re- gions were tested using dual luciferase kits. [Result]One transcription starting sites（TSS）, two CpG islands and multiple transcription factor binding s |
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| Publisher | 广西大学 动物科学技术学院,南宁,530004 |
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| Snippet | 【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... S828.89; [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... |
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| SubjectTerms | ANK1基因 启动子 广西巴马小型猪 活性分析 锚蛋白（ANK） |
| Title | 广西巴马小型猪ANK1基因启动子克隆及其活性分析 |
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