努比亚山羊UCP 1基因克隆、生物信息学分析及组织表达研究  

Cloning,Bioinformatics Analysis and Tissue Expression of UCP 1 Gene in Nubian Goats

作　　者：韦义荣 郑自华[2] 张叁保 宋颖 蒋海玉 孙雯玥 程鹏飞 刘雨帆 邹剑伟 黄艳娜[1] 潘艳 蒋钦杨[1] WEI Yirong;ZHENG Zihua;ZHANG Sanbao;SONG Ying;JIANG Haiyu;SUN Wenyue;CHENG Pengfei;LIU Yufan;ZOU Jianwei;HUANG Yanna;PAN Yan;JIANG Qinyang(College of Animal Science and Technology,Guangxi University,Nanning 530004,China;Agricultural and Animal Husbandry Industry Development Research Institute,Guangxi University,Nanning 530004,China;Guangxi Vocational University of Agriculture,Nanning 530009,China)

机构地区：[1]广西大学动物科学技术学院,南宁530004 [2]广西大学农牧产业发展研究院,南宁530004 [3]广西农业职业技术大学,南宁530009

出　　处：《中国畜牧兽医》2023年第2期440-450,共11页China Animal Husbandry & Veterinary Medicine

基　　金：国家重点研发计划项目(2021YFD1100100);国家现代农业产业体系广西牛羊产业创新团队建设项目(nycytxgxcxtd-2021-09);广西重点研发计划项目(桂科AB20297012)。

摘　　要：【目的】试验旨在克隆努比亚山羊解偶联蛋白-1(uncoupling protein-1,UCP1)基因并进行生物信息学分析,检测其在努比亚山羊不同组织中的表达差异,为研究努比亚山羊UCP 1基因功能及进一步解析其在脂肪代谢中的调节作用提供数据。【方法】以努比亚山羊皮下脂肪组织cDNA为模板,采用PCR扩增并克隆UCP 1基因CDS区序列后,与其他物种进行相似性比对及系统进化树构建,并对UCP1蛋白进行生物信息学分析;利用实时荧光定量PCR方法检测UCP 1基因在努比亚山羊心脏、肝脏、脾脏、肾脏、背最长肌、皮下脂肪、腹脂中的相对表达量。【结果】努比亚山羊UCP 1基因CDS区全长918 bp,编码305个氨基酸。相似性比对发现,努比亚山羊UCP 1基因氨基酸序列与绵羊、瘤牛×普通牛、水牛、羚羊、马鹿、双峰驼、驴、大熊猫、人的相似性分别为98.1%、97.0%、96.5%、96.1%、95.8%、91.0%、87.0%、86.5%和83.8%。系统进化树表明,努比亚山羊与绵羊亲缘关系最近,与人的亲缘关系最远。生物信息学分析表明,努比亚山羊UCP1蛋白的分子质量为32.97 ku,等电点为9.29,属于碱性蛋白;UCP1蛋白缺乏稳定性,总平均亲水性为0.20,具备一定亲水性;脂溶性系数为91.70,跨膜螺旋氨基酸残基数量的预测值为15.94,前60个氨基酸的跨膜螺旋数预测值为8.76,位于膜细胞质侧的总概率为29.03%,属于非分泌蛋白;UCP1蛋白二级结构由α-螺旋(48.85%)、无规则卷曲(27.54%)、延伸链(16.39%)、β-转角(7.21%)构成;UCP1蛋白含有28个磷酸化位点、5个O-糖基化位点及2个N-糖基化潜在位点。实时荧光定量PCR结果显示,UCP 1基因在努比亚山羊皮下脂肪中表达量最高,显著高于其他组织(P<0.05);在背最长肌中表达量最低。【结论】试验成功扩增UCP 1基因CDS区序列,UCP1是一个缺乏稳定性、亲水的碱性蛋白;UCP 1基因在努比亚山羊各组织中均有表达,且主要在皮下脂肪及脾�【Objective】This study was aimed to clone uncoupling protein-1(UCP1)gene in Nubian goats and carried out bioinformatics analysis,and detect its expression difference of UCP 1 gene in different tissues of Nubian goats,in order to provide data for studying its regulatory function in fat metabolism and analyzing the function of UCP 1 gene in Nubian goats.【Method】Using the cDNA from the subcutaneous fat tissue in Nubian goats as a template,the CDS region of UCP 1 gene in Nubian goats was amplified by PCR and cloned,sequence similarity alignment and phylogentic tree contruction were carried out with other species,and the bioinformatics analysis of UCP1 protein was carried out.The expression of UCP 1 gene in heart,liver,spleen,kidney,longissimus dorsi muscle,subcutaneous fat and abdominal fat of Nubian goats were detected by Real-time quantitative PCR.【Result】The CDS sequence of UCP 1 gene in Nubian goats was 918 bp and encoded 305 amino acids.The similarity of amino acid sequence of UCP 1 gene between Nubian goats and Ovis aries,Bos indicus×Bos taurus,Bubalus bubalis,Oryx dammah,Cervus elaphus,Canelus ferus,Equus asinus,Aliuropoda melanoleuca and Homo sapiens were 98.1%,97.0%,96.5%,96.1%,95.8%,91.0%,87.0%,86.5%and 83.8%,respectively.The phylogenetic tree indicated that the closest was to Ovis aries and the furthest was to Homo sapiens.The results of bioinformatics analysis showed that the molecular mass of UCP1 protein in Nubian goats was 32.97 ku,with an isoelectric point of 9.29,which was a basic proteins.UCP1 protein lacked stability,an overall average hydrophilicity was 0.20,which possessed certain hydrophilicity.The lipid solubility coefficient was 91.70,the predicted value of the number of amino acid residues in transmembrane helices was 15.94,the predicted value of the number of transmembrane helices in the first 60 amino acids of the protein was 8.76,the overall probability of being located on the cytoplasmic side of the membrane was 29.03%,and it belonged to nonsecretory proteins.The secondary str

关 键 词：努比亚山羊 UCP 1基因 克隆 生物信息学分析 组织表达 

分 类 号：Q78[生物学—分子生物学]