Genomic Imprinting Of Dio3、Dio3os 、Wif1 And The Dna Methylation Status Of Wif1 Gene In Cattle

Genomic imprinting can lead to the phenomenon of differential expression of the parental alleles depending on DNA or histone alleles by epigenetic modification. The gene which paternal allele silent, while the maternal allele activated gene called paternally imprinted gene; conversely, called maternal imprinted genes. Currently, the number of imprinted genes in human and mice have been identified is more than 200, but relatively few studies in bovine.This study selected Dlk1-Dio3 imprinted domain in mice identified as paternal expression Dio3, Dio3 os and Wif1 gene imprinted in human placenta. By RT-PCR method,the expression status of the three genes was analyzed in various tissues of adult holstein cattles. The results showed that the three genes in adult bovine heart, liver, spleen, lung, kidney, muscle and fat have expressed into RNA. In this study, allelic expression analysis was based on SNP(single nucleotide polymorphism, SNP) sites sequencing. First, the method of sequencing the PCR products was used to find SNP sits of Dio3, Dio3 os and Wif1 genes to identify the heterozygotes. By directly sequencing the RT-PCR products of three genes in the heterozygous, we can find that Dio3 and Dio3 os were showed in adult cattle tissues and Wif1 gene was showed monoallelic expression in the lungs, while showed biallelic expression in heart, liver, spleen, kidney, muscle, and fat.To analyze the possible role of DNA methylation in regulating Wif1 tissue-specific imprinting, the methylation status of 29 Cp G sites in Wif1 gene promoter region was analyzed in lungs(monoallelic expressed tissues) and livers(biallelic expressed tissue) by bisulfite sequencing. The results showed that the methylation levels between the two parental strands exhibited no significant difference(P> 0.05) neither in lung nor liver tissues. The difference of methylated rates of each Cp G site between two parental chains in lungs and livers was further analyzed. In lungs, there are 8 Cp G sites which exhibies significant differences of methylation rate between parental chains compared to those in livers. Among them, Cp G sites 1、2 and 6 were contained in the transcription factor biding site. Because the methylation changes of individual Cp G sites can effect the binding of transcription factors, the methylation of these three Cp G sites in Wif1 promoter region may be involved in the regulation of tissue-specific imprinting of cattle Wif1 gene.