奶山羊短链脂肪酸受体基因(GPR43)编码区(CDS)的克隆及表达分析  

Coding Sequence(CDS) Cloning and Expression Analysis of Short-chain Fatty Acid Receptor Gene(GPR43) in Dairy Goat(Caprahircus)

作　　者：孙雨婷[1] 罗军[1] 邱思源 田慧彬 

机构地区：[1]西北农林科技大学动物科技学院陕西省农业分子生物学重点实验室,杨凌712100

出　　处：《农业生物技术学报》2014年第7期876-882,共7页Journal of Agricultural Biotechnology

基　　金：转基因生物新品种培育项目(No.2009ZX08009-162B);陕西省重大科技创新项目(No.2009ZKC07-01-1);公益性行业(农业)科研专项(No.201103038)

摘　　要：GPR43是G蛋白偶联受体(G protein-coupled receptors,GPCRs)家族成员之一,在脂代谢中具有潜在的调控作用。本实验旨在克隆奶山羊(Capra hircus)短链脂肪酸受体基因GPR43编码区(coding sequence,CDS)并分析其组织表达谱。根据GenBank上已登录的牛(Bos taurus)GPR43基因(NM_001163784)序列设计引物,利用RT-PCR方法克隆奶山羊GPR43基因的编码区(coding sequence,CDS)(GenBank登录号:HM623658),测序结果分析表明,该基因CDS区长度为987 bp,共编码329个氨基酸。序列同源性分析表明,奶山羊GPR43的核苷酸和氨基酸序列与牛、人(Homo sapiens)、鼠(Mus musculus)的相似性较高,且与牛的相似性高达90%以上。蛋白结构预测发现,奶山羊GPR43蛋白具有7个跨膜螺旋,且C端存在较强的疏水区域,N端则表现出较强的亲水性。实时定量PCR分析表明,在干奶期奶山羊的10个组织中,GPR43基因在脾脏中表达量最高,小肠、肝脏、脂肪次之,肺脏、肌肉和肾脏表达相对最少,而乳腺中也有少量的表达,表明其具有明显的组织表达特异性。泌乳期奶山羊GPR43基因的表达显著高于干奶期,表明该基因在奶山羊泌乳组织中具有一定的调控作用。研究结果为进一步揭示GPR43在奶山羊乳腺组织的功能提供参考资料。G protein-coupled receptor 43（GPR43） is the member of G protein-coupled receptors（GPCRs） family and has a potential role of regulating lipid metabolism.The aim of this study was to clone GPR43 and analyze its mRNA expression level in different tissues and lactation periods of diary goats（Capra hircus）.Based on the published nucleotide sequence of bovine（Bos taurus） GPR43 gene（NM_001163784） in GenBank,a pair of specificity primers was designed.Coding sequence（CDS） of GPR43 was amplified （GenBank accession number：HM623658） by RT-PCR.The results showed that the CDS length of GPR43 gene was 987 bp,coding 329 amino acids.The homology analysis of dairy goat GPR43 gene indicated that the nucleotides and amino acids had high similarity with bovine,human（Homo sapiens） and mice （Mus musculus） and reached up to more than 90％ with bovine.The protein structure analysis showed that GPR43 protein had 7 transmembrane helices and the C-terminal was high hydrophobic but the N-terminal showed high hydrophilism.Real-time quantitative PCR was used to detect the expression of GPR43 gene in different tissues.The results suggested that GPR43 mRNA expressed in all 10 tissues examined.GPR43 mRNA expressed abundantly in spleen,followed by the small intestine,liver,adipose tissues but the lowest in lung,muscle and kidney,and the relative low expression was also detected in mammary gland tissue.Two lactation stages expression analysis showed that expression of GPR43 gene was significantly higher in the lactation stage than that of dry stage （P＜0.05）.Taken together,these results demonstrated that GPR43 had a different range of tissue-specific expression,and might play an important role during lactation stages in dairy goat.In conclusion,this work serves as the basis for further study on GPR43 in the dairy goat mammary gland.

关 键 词：奶山羊 G protein-coupled receptor 43(GPR43) 克隆 表达分析 

分 类 号：Q343.1[生物学—遗传学]