西农萨能奶山羊LXRα基因的克隆、序列分析及组织表达

采集西农萨能奶山羊乳腺组织,利用RT-PCR和RACE技术分离并克隆LXRα基因的cDNA序列。其全长1 654 bp(GenBank收录号为GU332719),包括5′UTR150 bp,CDS1 344 bp和3′UTR160 bp,该基因编码447个氨基酸。经氨基酸序列分析发现,山羊LXRα与GenBank中牛(Bos taurus)、小鼠(Mus muscu-lus)、褐鼠(Rattus norvegicus)、猪(Sus scrofa)和人(Homosapiens)的LXRα相似性较高,均在90%以上。蛋白质结构分析表明,其蛋白质分子量为50.35 ku,等电点为6.3。具有受体特征结构域:配体结合区域(Lig-and-binding Domain,LBD)、DNA结合区域(DNA-binding Domain,DBD)和锌指蛋白C4型区域(Zinc fingerC4 type,zf-C4)。整个序列具有较强的亲水性且不含信号肽与跨膜结构,该基因定位于细胞核。此外,还通过实时荧光定量PCR(RT-qPCR)技术对LXRα基因进行组织表达分析。在所检测的西农萨能奶山羊11个组织中均有LXRα基因的mRNA存在,其中小肠组织中表达最丰富,乳腺、肝脏、脾脏、皮脂次之,心脏相对最低。

Cloning,Sequence Analysis and Tissues Expression Research of LXRa Gene on Xinong Saanen Dairy Goats

LXRα(Liver x receptor α)is a key sensor of lipid homeostasis.This experiment was conducted to clone LXRα gene cDNA sequence,to analyze the sequence by biology information analyzing software,and to study the gene expression in differences tissues.In this experiment,RT-PCR and RACE methods were used for the isolation and cloning of the whole cDNA in mammary gland.The length with 1 654 bp(GenBank accession No.GU332719) which contain 150 bp 5′UTR,1 344 bp CDS and 160 bp 3′UTR,encoding a protein of 477 amino acid residues.The amino acid of Capra hircus LXRα with high score similarity,more than 90%,compared to Bos taurus,Mus musculus,Rattus norvegicus,Sus scrofa and Homo sapiens in GenBank which was indicated by amino acid sequence alignment.The protein structure analysis showed that the protein molecular weight is 50.35 ku and the isoelectric point is 6.3.It has the characteristic of receptor domain: Ligand-binding Domain(LBD),DNA-binding Domain(DBD) and Zinc finger C4 type(zf-C4).The whole sequence has strong hydrophilic,does not have signal peptide and transmembrane structure,localized in the nucleus.In addition,we also analyzed the tissue expression of LXRα gene through the approach of real-time fluorescence quantitative-PCR(RT-qPCR).The results showed that the mRNA of LXRα gene existed in all the 11 detected organizations with the most abundant expression in small intestine tissue,followed by the mammary gland,liver,spleen and sebum,the minimal expression in heart was observed.