Study Of Influence Of Mutation Of Foxp3 Binding Site In Bic On Expression Of MiR-155 In Pig

In recent years, resistant breeding has been highly concerned by home and abroad researchers in the areas of agricultural animal breeding and genetics. China is an important parental source of local breeds in resistance breeding selection, so analysis of the immunity differences between Chinese local breeds and exotic breeds is necessary to analyze whether there is stronger resistance of local breeds compared with exotic breeds and breed new resistance breeds from the perspective of heredity. Mi R-155 plays an essential role in the development, occurrence and regulation of immune system, and their differential expression in different breeds may lead to differences in the entire immune system. This study aims to use chromatin immunoprecipitation technology to research regions of swine Bic gene binded with Foxp3 protein, analyze the binding differences between regions with different A/C mutations founded in the Bic gene of Large White and Meishan pigs with transcription factors Foxp3 by the dual luciferase reporter system,and detect the expression level of mi R-155 and its’ candidate target genes at the individual level by quantitative PCR, and validate the expression levels of a candidate target gene of mi R-155 with dual fluorescence experiments. Concrete results obtained are as follows:(1) Based on the 5 SNP obtained in our laboratory, a fragment of approximately 3kb was amplified before and after pre-mi R-155 in Large White and Meishan pigs respectively, and obtained 20 suspected SNP loci by sequencing.(2) Initially identified that transcription factor Foxp3 in pig could bind with AAACA motif in about 305 bp upstream of pre-mi R-155.(3) The existence of SNP in different breeds leaded different binding abilities of correspondent motif with Foxp3, and binding abilities of AAACC motif with Foxp3 was stronger than AAACA motif. Furthermore, this combination may require certain conditions such as fragment length or the number of motifs.(4) The different binding abilities of transcription factor Foxp3 with two haplotypes affected the expression of mi R-155 in pigs, and the expression level of mi R-155-5p and mi R-155-3p in individuals with CC genotype was higher than it with AA genotype.(5) Preliminary determinated that the expression of candidate target genes of mi R-155 was higher in individuals with AA genotype compared with individuals with CC genotype, and identified the targeted relationship between mi R-155 and Sept11, Phf17 genes.This study proved the changes of mi R-155 and its target genes due to the different locus binded with Foxp3 in Bic gene of Large White and Meishan pigs, provided some theoretical basis for revealing immunity differences between these two breeds, as well as supply new molecular identification basis for new resistance breeds selection.