猪A-FABP基因真核表达载体的构建及其组织表达

Constructing of Eukaryotic Expressing Vector and Tissues Expression of A-FABP Gene in Pig

本实验以长白猪背最长肌为实验材料,进行组织RNA的提取及A-FABP基因的克隆,构建p EGFPN1-A-FABP真核表达载体,并通过脂质体转染成纤维细胞,48 h观察荧光表达。同时应用荧光定量PCR法检测猪7种不同组织(心脏、肝脏、脾脏、肺脏、肾脏、背最长肌、腿肌)中A-FABP基因的差异表达。结果表明:成功构建p EGFP-N1-A-FABP融合表达载体,并在细胞中表达绿色荧光蛋白;A-FABP基因在7种组织中均有表达,其中背最长肌和腿肌中A-FABP基因的表达量最高,与其他组织相比差异极显著(P<0.01),而脾脏和肾脏中的表达量相对于心脏、肝脏和肺脏差异显著(P<0.05),而A-FABP基因表达量在心脏、肝脏和肺脏中差异不显著(P>0.05)。表明该基因能够在真核载体和细胞中表达,并且在猪不同组织的表达具有差异性。

The longissimus dorsi muscle of Landrace pig was as experimental material, RNA was extracted and A-FABP gene was cloned, the eukaryotic expression vector of pEGFP-N1-A-FABP was constructed and transfected to fibroblasts by liposome, the fluorescence expression was detected after 48 h. Then the differential expression of A-FABP gene in different tissues （heart, liver, spleen, lung, kidney, longissimus dorsi muscle and leg muscle） was detected by Q-PCR. Results showed that the pEGFP-N1-A-FABP fusion expression vector was successfully constructed, the fluorescence expression was successfully expressed in fibroblasts. The expression ofA-FABP gene in 7 different tissues was different. The gene expression in longissimus dorsi muscle and leg muscle were the highest compared with others, they had extremely significant difference （P〈0.01）, the expression in spleen and kidney were significantly different from that of heart, liver and lung （P〈0.05）, while the A-FABP gene was not significantly different in heart, liver and lung. Results indicated that A-FABP gene was expressed in eukaryotic vector and cells, and its expression had difference in different tissues oldies.