三角鲂MHCⅠα基因全长cDNA克隆与生物信息学分析  

Full-length cDNA cloning and bioinformatic analysis of MHCⅠαgene from black amur bream(Megalobrama terminalis)

作　　者：刘凯[1] 谢楠[1] 郭炜 马恒甲[1] LIU Kai;XIE Nan;GUO Wei;MA Hengjia(Institute of Fishery Science,Hangzhou Academy of Agricultural Sciences, Hangzhou 310024, China)

机构地区：[1]杭州市农业科学研究院水产研究所,浙江杭州310024

出　　处：《浙江农业学报》2022年第6期1162-1174,共13页Acta Agriculturae Zhejiangensis

基　　金：杭州市农科院创新基金(2019HNCT-01);杭州市农业科研主动设计项目(20162012A03);现代农业产业技术体系专项(CARS-45-39)。

摘　　要：主要组织相容复合体(major histocompatibility complex,MHC)是一类具有高度多态性的分子,在脊椎动物的先天免疫应答中起到重要作用。根据已报道鱼类的MHCⅠα基因序列设计特异性引物,通过RT-PCR和RACE技术克隆三角鲂MHCⅠα基因cDNA序列全长,命名为Mete-UAA,并对其进行生物信息学分析。结果显示,Mete-UAA全长为2102 bp,包含1044 bp的开放阅读框,编码347个氨基酸。编码的MHCⅠα蛋白质预测分子量为38699.18,等电点5.23,含有16个氨基酸组成的信号肽,具有亲水性,定位于细胞膜上,具有1个N-糖基化、3个O-糖基化和39个磷酸化的潜在位点。氨基酸序列同源性比对发现,Mete-UAA氨基酸序列与团头鲂同源性最高为75.79%,与草鱼同源性稍低于团头鲂,为75.25%。Mete-UAA具有MHCⅠα的典型结构特征,包含1个前导肽、3个胞外结构域、1个跨膜区和1个胞质区。二级结构分析显示,Mete-UAA蛋白质中α-螺旋、β-转角、延伸链和无规则卷曲所占的比例分别为25.94%、9.22%、26.80%和38.04%。基于转录组学分析表明,感染嗜水气单胞菌(Aeromonas hydrophila)后三角鲂MHCⅠα基因表达呈总体下降趋势。本研究从三角鲂肝中成功克隆MHCⅠα基因,分析了其生物信息学特征,为后续研究MHCⅠα参与调控三角鲂先天免疫应答的机制提供理论基础。Major histocompatibility complex(MHC)is a highly polymorphic molecule that plays an essential role in vertebrates'innate immune response.Specific primers were designed according to the reported MHCⅠαsequence of fish.Finally,the full-length MHCⅠαcDNA sequence of Megalobrama terminalis was obtained by RT-PCR and RACE.The cDNA sequence was named Mete-UAA and conducted bioinformatics analysis.The results showed that Mete-UAA was 2102 bp in length,open reading frame was 1044 bp,encoding 347 amino acids.The Mete-UAA encoded a hydrophilic protein that contained 16 amino acid signal peptides.It was localized on the cell membrane and had one N-glycosylation,three O-glycosylation,and 39 potential phosphorylation sites.The encoded MHCⅠαprotein had a predicted relative molecular mass of 38699.18 and an isoelectric point of 5.23.Amino acid sequence homology comparison showed that the homology of the Mete-UAA amino acid sequence was up to 75.79%with M.amblycephala(AEE87250),and 75.25%with Ctenopharyngodon idella(BAD01521),slightly lower than M.amblycephala(AEE87250).The Mete-UAA possessed the typical structural features of MHCⅠα,including a leader peptide,3 extracellular domains,1 transmembrane region,and 1 cytoplasmic region.The secondary structure analysis of the Mete-UAA protein showed that the proportions of theα-helix,β-turn,extended chain and random coil were 25.94%,9.22%,26.80%,and 38.04%,respectively.Based on the transcriptome analysis,expression of the MHCⅠαgene in M.terminalis infected with Aeromonas hydrophila showed an overall decreasing trend.In this study,the MHCⅠαgene was successfully cloned from the liver of M.terminalis.Its bioinformatics characteristics were analyzed,which would provide a theoretical basis for further research on the mechanism of MHCⅠαinvolved in regulating the innate immune response in M.terminalis.

关 键 词：三角鲂 主要组织相容复合体 基因克隆 生物信息学分析 

分 类 号：S917.4[农业科学—水产科学]