广灵驴DGAT 2基因克隆、生物信息学分析及组织表达研究  

Cloning,Bioinformatics and Tissue Expression Analysis of DGAT2 Gene in Guangling Donkey

作　　者：李武峰[1] 关家伟 孙瑜彤 邱丽霞 杜敏 LI Wufeng;GUAN Jiawei;SUN Yutong;QIU Lixia;DU Min(College of Life Sciences,Shanxi Agricultural University,Taigu 030801,China;Department of Animal Sciences,Washington State University,Pullman 99164-6310,USA)

机构地区：[1]山西农业大学生命科学学院,太谷030801 [2]美国华盛顿州立大学动物科学系,普尔曼99164-6310

出　　处：《中国畜牧兽医》2021年第2期407-416,共10页China Animal Husbandry & Veterinary Medicine

基　　金：山西省重点研发计划(指南)项目:国际科技合作方面(201803D421022);山西农业大学横向科技项目(2019HX78)。

摘　　要：本研究旨在对广灵驴二酰基甘油酰基转移酶2(DGAT2)基因进行克隆,生物信息学分析和检测其在不同组织中的表达情况,为探究DGAT2基因在广灵驴脂肪沉积和提高乳脂率等方面的作用机制提供理论参考。根据GenBank上公布的马(登录号:XM_023645689.1)、双峰驼(登录号:XM_010973154.1)、绵羊(登录号:XM_027979550.1)等物种的DGAT 2基因mRNA序列,利用Primer Premier 3.0在线工具设计同源引物,应用RT-PCR法扩增DGAT 2基因序列,用生物信息学方法分析DGAT 2基因编码序列,用实时荧光定量PCR技术检测DGAT 2基因在广灵驴心脏、肝脏、脾脏、肺脏、肾脏、背最长肌、肌间脂肪、皮下脂肪组织中的表达。结果显示,广灵驴DGAT 2基因CDS序列1086 bp,编码361个氨基酸,提交到GenBank,获得登录号:MT993643,其编码序列与马、牛、双峰驼、猪、绵羊、人、小鼠的同源性分别为99.0%、92.0%、93.5%、92.0%、92.7%、85.3%、84.1%。系统进化树分析表明,驴与马的亲缘关系最近,和小鼠的关系最远。DGAT2蛋白分子质量40.96 ku,脂肪系数92.85,等电点9.16,是一种具有跨膜区的稳定碱性疏水蛋白。DGAT2蛋白有28个磷酸化修饰位点,2个糖基化修饰位点,没有信号肽,主要定位在内质网,α-螺旋(39.89%)和无规则卷曲(36.01%)是主要的二级结构。DGAT 2基因在检测的8个组织中都有表达,其中皮下脂肪中的表达量显著高于其余组织(P<0.05),其次是心脏、肝脏和肾脏,背最长肌中的表达量最低。本试验结果为探究DGAT 2基因在广灵驴脂肪沉积和提高乳品质性状的作用奠定基础。The study was aimed to clone,bioinformatics of diacylglycerol acyltransferase 2(DGAT2)gene of Guangling donkey,detected its expression in different tissues,and provided theoretical reference for exploring the mechanism of DGAT2 gene in fat deposition and improving milk fat rate of Guangling donkey.Homologous primers was designed by Primer Premier 3.0 according to the mRNA sequences of DGAT2 gene of Equus caballus(accession No.:XM_023645689.1),Camelus bactrianus(accession No.:XM_010973154.1),Ovis aries(accession No.:XM_027979550.1),and other species published in GenBank.DGAT2 gene sequence of Guangling donkey was amplified with RT-PCR.The encoding sequence of DGAT2 gene was analyzed by bioinformatics methods,and the expression of DGAT2 gene in heart,liver,spleen,lung,kidney,longissimus dorsi muscle,intermuscular fat and subcutaneous fat of Guangling donkey was detected by Real-time quantitative PCR.The results showed that the CDS of Guangling donkey DGAT2 gene consisted of nucleotides of 1086 bp encoding 361 amino acids.It was submitted to NCBI and the login number was MT993643.The coding sequence of DGAT2 gene showed 99.0%,92.0%,93.5%,92.0%,92.7%,85.3%and 84.1%identity with that of Equus caballus,Bos Taurus,Camelus bactrianus,Sus scrofa,Ovis aries,Homo sapiens and Mus musculus.Phylogenetic tree analysis revealed that Guangling donkey was most closely related to Equus caballus and farthest related to Mus musculus.The DGAT2 protein,with molecular weight of 40.96 ku,fat coefficient of 92.85 and isoelectric point of 9.16,was a stable basic hydrophobic protein with transmembrane region.There were 28 phosphorylation modification sites and 2 glycosylation modification sites in DGAT2 protein,with no signal peptide,it was mainly located in the endoplasmic reticulum.The secondary structure of DGAT2 protein was mainly of 39.89%alpha helix and 36.01%random coil.The DGAT2 gene was expressed in 8 tissues,among which the expression level in subcutaneous fat was significantly higher than that in other tissues(P<0.05),followed by

关 键 词：DGAT 2基因 广灵驴 基因克隆 生物信息学分析 组织表达 

分 类 号：S822[农业科学—畜牧学]