生长激素受体基因突变对西藏藏猪生长迟缓的影响  

The Impact of GHR Mutation on Suppressing the Growth in Tibet Mini-pig

作　　者：岳敏[1] 田雨光[1] 万斌[1] 庞炜[1] 吴清洪[1] 王玉珏[1] 

机构地区：[1]南方医科大学实验动物中心,广州510515

出　　处：《畜牧兽医学报》2016年第5期882-887,共6页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金(801402625);中国博士后自然科学基金(2014M550439);广东省自然科学基金(S2013010014720);广东省科技计划项目(2011B060300014);南方医科大学2015科研助手项目(C1032246)

摘　　要：本研究旨在探索西藏藏猪生长迟缓成因和分子机理。采用逆转录聚合酶链反应(RT-PCR)技术对西藏藏猪生长激素受体(GHR)基因克隆,采用生物信息学方法分析西藏藏猪生长激素受体基因与其他常见品种猪基因与蛋白序列差异,并构建GHR基因的真核过表达质粒,转染到西藏藏猪胚胎成纤维细胞(PEFs)上进行瞬时表达,通过MTT检测细胞增殖活性,利用实时荧光定量PCR技术检测细胞的胰岛素样生长因子-1(IGF-1)表达的情况。西藏藏猪GHR基因编码区1 716bp,编码572个氨基酸;与普通猪GHR基因序列进行比对分析表明,在西藏藏猪GHR基因编码区1 225bp处发生T→G突变,导致丝氨酸突变为丙氨酸;GHR基因真核过表达质粒成功转染PEFs后,PEFs生长活性显著提高;qPCR检测表明,细胞株中IGF-1表达显著上调。综上表明,GHR基因的表达可以提高PEFs细胞的增殖活性,诱导IGF-1基因的表达,GHR基因在1 225bp处的点突变可能影响西藏藏猪的生长迟缓。This study aimed to explore the genetic and molecular mechanism of Tibet mini-pig growth retardation. RT-PCR technique was used to amplify Tibet mini-pig GHR gene. Bioinfor- matics method was used to analyze the differences of GHR gene and protein sequence between Ti- bet mini-pig and other common variety pigs. The eukaryotic expression vector of common pig GHR gene was transfected into Tibet mini-pig＇s embryonic fibroblasts. The MTT assay was used to detect the cell proliferation activity after transfected GHR-pIRES2-EGFP vector. The real-time PCR was used to detect the mRNA expression of the IGF-1 gene. There were 1 716 bp in Tibet mini-pig GHR gene coding region, and 572 amino acids were encoded. Compared with common meat pig GHR gene sequence,the base T in Tibet mini-pig GHR gene coding region 1 225 bp mu- tated into G,and then serine was replaced by alanine. The growth activity of Tibet mini-pig＇s em- bryonic fibroblasts were significantly improved after transfected with GHR-pIRES2-EGFP, and the mRNA level of IGF-1 was up-regulated significantly. The results showed that GHR could im- prove the activity of PEFs cell proliferation,and induce the expression of IGF-1 gene;The muta- tion of GHR gene in 1 225 bp might involve in the process of growth retardation in Tibet mini-pig.

关 键 词：西藏藏猪 GHR IGF-1 胚胎成纤维细胞 

分 类 号：S828.2[农业科学—畜牧学]