FUT1基因启动子区对断奶仔猪大肠杆菌抗性的调控机制分析

项目名称： FUT1基因启动子区对断奶仔猪大肠杆菌抗性的调控机制分析

项目编号： No.31472066

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 畜牧学与草地科学

项目作者： 吴圣龙

作者单位： 扬州大学

项目金额： 90万元

中文摘要： 大量研究表明，FUT1基因在断奶仔猪抗大肠杆菌感染过程中发挥了重要的调控作用，但是中国地方猪种FUT1基因编码区高度保守，因此本研究锁定其启动子区，通过生物信息学预测和实验验证确定核心启动子序列，在确证的梅山猪抗性和敏感型断奶仔猪中，检测核心启动子区域SNPs及甲基化位点，结合转录因子结合位点的预测分析，筛选顺式作用元件中重要的变异位点，系统分析其与断奶仔猪大肠杆菌抗性的关系；同时，利用生物信息学、文献挖掘、点突变分析、EMSA和CHIP等手段鉴定关键的转录因子及顺式作用元件，在猪小肠上皮细胞系中过表达和干扰（基因敲入）关键的转录因子，分析验证关键的转录因子及顺式作用元件对基因表达及断奶仔猪大肠杆菌抗性的影响；通过对重要变异位点、关键转录因子及顺式作用元件功能的系统分析和层层验证，以期阐明FUT1基因启动子区对断奶仔猪大肠杆菌抗性的作用及其机制，为地方猪种抗大肠杆菌病育种提供依据和指导。

中文关键词： 分子机制；抗病育种；表达调控；DNA甲基化

英文摘要： A large number of studies have shown that α(1, 2)-fucosyltransferase gene (FUT1) plays an important role in regulating the process of the resistance to Escherichia coli infection in weaning piglets, but the coding region of FUT1 gene shows highly conservative in Chinese native pig breeds. Therefore this study focuses on the analysis of FUT1 promoter region and confirms the key promoter sequence by bioinformatics prediction and experimental verification. Additionally, the SNPs (single nucleotide polymorphisms) and methylation sites of FUT1 key promoter region were detection in the selected individuals of E.coli F18-resistant and -sensitive Meishan weaning piglets. The important variation sites were screened out combining the predictive analysis of transcription factor binding sites and systematically analyzed whether the variation sites were related to the resistance to E.coli of weaning piglets. Meanwhile the key transcription factors and cis-acting elements were identified with the methods of bioinformatics, literature mining, point mutation analysis, EMSA (Electrophoretic Mobility Shift Assay) and CHIP (Chromatin Immunoprecipitation). Using the overexpression and interference experiment of key transcription factors in pig small intestine epithelium, and then analyzed and validated whether key transcription factors or cis-acting elements affected the expression level of FUT1 gene and the resistance to E.coli of weaning piglets. Through the systematic analysis and continuous verification of the function of key transcription factors and cis-acting elements, which aimed to illuminate the regulation mechanism of resistance to E.coli by FUT1 promoter region in weaning piglets, and lay a solid foundation for the breeding of disease resistance to E.coli in Chinese native pig breeds.

英文关键词： Molecular mechanism;Disease-resistance breeding;Expression regulation;DNA hypermethylation