苏太断奶仔猪TAP1基因启动子区甲基化修饰与F18大肠杆菌抗性的关系

Correlation Between TAP1 Gene Promoter Methylation Modification and the Resistance to E. coli F18 in Sutai Weaning Piglets

本试验运用亚硫酸氢盐(BSP)+Miseq测序法分析苏太断奶仔猪F18大肠杆菌抗性型和敏感型个体十二指肠和空肠组织TAP1基因启动子区的甲基化水平,并运用real-time PCR(RT-PCR)方法检测TAP1的m RNA表达量,进而分析TAP1基因启动子区甲基化修饰对m RNA表达的影响,探讨TAP1基因启动子区甲基化修饰对F18大肠杆菌抗性的调控作用。结果表明:TAP1基因启动子区2个Cp G岛及其中间区域内存在28个Cp G位点,且都存在不同程度的甲基化;其中在Cp G-6位点,抗性型个体空肠组织中的甲基化水平显著低于敏感型个体的甲基化水平(P<0.05);在Cp G-7和Cp G-8位点,抗性型个体十二指肠组织中的甲基化水平显著高于敏感型个体的甲基化水平(P<0.05);Cp G-7和Cp G-8位点甲基化水平与TAP1基因m RNA表达量呈负相关。本实验结果显示,Cp G-6、Cp G-7和Cp G-8是调控基因转录水平的关键性位点,断奶仔猪可能通过对TAP1基因启动子区Cp G岛这些关键位点的去甲基化来提高十二指肠和空肠组织中m RNA的表达水平,进而发挥对E.coli F18抗性的调控作用。

TAP1 gene promoter methylation levels in duodenum and jejunum tissues from Sutai resistant and sensitive weaning piglets were analyzed with Bisulfite Sequencing PCR （BSP） ＋ Miseq methods, meanwhile mRNA expression of TAP1 gene was detected by real-time PCR （RT-PCR）, and then the associations between the methylation level of TAP1 gene promoter and mRNA expression was analyzed in this the regulation mechanism of TAP1 gene methylation modification experiment. The aim of this study was to investigate on the resistance to E. coli F18. The results showed that in TAP1 gene promoter region there were 28 CpG sites between the two CpG islands and their middle area, and all the sites were in different degrees of methylation; At the CpG-6 site, the methylation level of resistant group in jejunum tissue was significantly lower than that of the sensitive group （P〈O.05）. At the CpG-7 and CpG-8 site, the methylation level of resistant group in duodenum tissue was significantly higher than that of the sensitive group （P〈 0.05）. The methylation levels of CpG-7 and CpG-8 site were negatively correlated with the expression of mRNA. The results indicated that CpG-6, CpG-7 and CpG-8 may be the key sites for the regulation of gene transcription, these key sites may regulate the mRNA expression through the methylation level in the duodenum and jejunum tissue of weaning piglets, and plays an important role in regulation of E. coli F18 resistance.