关键词: 广西巴马小型猪, α-1, 3半乳糖转移酶, 真核表达载体, 表达鉴定

Abstract: Objective For the purpose of producing GGTA1 gene silencing bama-mini pig, cloned and analyzed the function of α-1,3 galactosyltransferase gene of bama-mini pig then constructed two expression vectors. Methods The GGTA1 gene was amplified from liver tissue by RT-PCR using a specific pair of primers which were designed and synthesized according to the GGTA1gene sequence released in GenBank, the amplified cDNA fragment was ligated into pMD18-T vector after purification. The recombinant was verified by the method of PCR and sequencing, then the enzyme digested products was ligated into pEGFP-N₁ to construct the expression vectors. identified by PCR and digesting reaction. Results The α-1,3 galactosyltransferase gene of bama-mini pig exist the absence of 5-exon(81-116) which containing 1080 bp, and existence of 5-exon which containing 1116 bp. Conclusions There exit the absence of 5-exon in the bama-mini pig, Construction of the both expression vectors (pEGFP-GGTA1-1 and pEGFP-GGTA1-2) can be transfected into PK-15 cells to identify their functional expression, also can be used in the experiment of gene silencing pig α-1,3 galactosyltransferase gene.

Key words: Bama-minipig in Guangxi, α-1, 3galactosyltransferase gene, Eukaryotic expression vector, Expression identification