GDF-9基因真核表达载体的构建及其对牛卵丘细胞扩展相关基因表达的影响

为构建牛生长分化因子9(GDF-9)基因真核表达载体,观察GDF-9基因真核表达载体转染牛卵丘细胞后对卵丘扩展相关基因表达的影响,试验在质粒pcDNA4 myc-his的多克隆位点插入GDF-9基因构建真核表达载体pcDNA4 myc-his-GDF-9,用脂质体转染牛卵丘细胞,利用Western blotting检测了GDF-9蛋白的表达量,同时利用实时荧光定量PCR(qRT-PCR)技术检测牛卵丘细胞中卵丘扩展相关基因的表达量.结果发现,转染了pcDNA4 myc-his-GDF-9的牛卵丘细胞中检测到GDF-9蛋白的表达,且牛卵丘细胞扩展相关基因PTGS2、PTX3和HAS2的表达量显著提高(P< 0.05),表明牛pcDNA4 myc-his-GDF-9可有效地转染到牛卵丘细胞中,该试验结果为进一步研究GDF-9基因的功能奠定了基础.

Construction of Eukaryotic Expression Vector of GDF-9 Gene and its Effect on the Expression of Expansion-related Genes in Bovine Cumulus Cells

The objective of the study was to construct the eukaryotic expression vector of bovine GDF-9 gene,and investigate its effect on the expression of expansion-related genes in transfected bovine cumulus cells.Eukaryotic expression vector plasmid pcDNA4 myc-his-GDF-9 was constructed by insert GDF-9 in the multiple cloning site.pcDNA4 myc-his-GDF-9 was transfected into bovine cumulus cells by liposome.The expression of GDF-9 gene was detected by Western blotting and the expression of the expansion-related genes in bovine cumulus cells was detected by qRT-PCR.The results showed that GDF-9 could express in transfected bovine cumulus cells and the expression of expansion-related genes PTGS2,PTX3 and HAS2 in transfected bovine cumulus cells were significantly increased (P< 0.05).In a conclusion,the bovine pcDNA4 myc-his-GDF-9 could effectively transfected into bovine cumulus cells,which could help to study farther functions of the GDF-9 gene.