关键词: 藏猪, DNA甲基转移酶3a基因, 单核苷酸多态性, 基因表达, 精子

Abstract: In order to screen the single nucleotide polymorphism( SNP) of DNA methyltransferase 3a(Dnmt3a) gene in Tibetan pig and analyze the expression level of this gene in Tibetan pig sperm,SNP of the gene was analyzed in Tibetan pig,Yarkshire pig and Diannan small-ear pig by PCR-direct sequencing,and it′s mRNA and protein expression levels were respectively measured in spermatozoa of Tibetan pig and Yarkshire pig by using real-time fluorescence quantitative PCR and Westen blot.The results showed that there were 2 SNPs in Dnmt3a gene 5′-flanking sequences and exon 11 region.One SNP(C→T) site was found at 2 012 bp upstream of the initial codon of Dnmt3a gene 5′-flanking sequences in Tibetan pigs,which had high heterozygosity and polymorphism information content,indicating there was large degree of variation within the population and more genetic information provided.However,this mutation site was not a peculiar SNP to Tibetan pig.The other SNP(C→T) site was found at 1 360 bp of exon regio.This SNP site had lower heterozygosity and polymorphic information content,indicating that the degree of variation within the population was smaller,and the genetic information provided was less.Both the mRNA and protein expression levels of Dnmt3a gene in spermatozoa of Tibetan pigs were significantly lower(P<0.05)than that of Yarkshire pig. The above results showed that the mutation site of 2 012 bp upstream of the initial codon in the 5′-flanking region of Dnmt3a gene in Tibetan pig could be used as a candidate marker in the molecular breeding.It might be one of the DNA methylation mechanism of Tibetan pig sperm plateau adaptation that there were lower mRNA and protein expression levels of Dnmt3a gene in spermatozoa of Tibetan pig compared to Yarkshire pig.

Key words: Tibet pig, DNA methyltransferase 3a gene, Single nucleotide polymorphism(SNP), Gene expression, Spermatozoa