凡纳滨对虾LvML1基因克隆与表达分析  

Molecular Cloning and Expression Analysis of LvML1 Gene in Pacific White Shrimp Litopenaeus vannamei

作　　者：刘洪涛 汪坤 杨明秋 LIU Hongtao;WANG Kun;YANG Mingqiu(Hainan Provincial Key Laboratory of Tropical Maricultural Technologies,Hainan Academy of Ocean and Fisheries Sciences,Haikou 571126,China)

机构地区：[1]海南省海洋与渔业科学院,海南省热带海水养殖技术重点实验室,海南海口571126

出　　处：《水产科学》2023年第2期175-184,共10页Fisheries Science

基　　金：财政部和农业农村部国家现代农业产业技术体系项目(CARS-48);海南省本级部门预算项目(2018-2019);海南省热带海水养殖技术重点实验室建设专项(2021)。

摘　　要：髓样分化蛋白2相关脂质识别(ML)蛋白是无脊椎动物先天免疫与脂质代谢方面具有重要功能的一类蛋白。克隆凡纳滨对虾1个新的ML基因cDNA全序列,命名为LvML1,包含254 bp的5′非编码区、913 bp的3′非编码区和462 bp的开放阅读框,开放阅读框共编码153个氨基酸。LvML1蛋白具有7个半胱氨酸残基,预测可形成3对二硫键,且具有1个典型ML家族结构域。多重序列对比发现,ML结构域和半胱氨酸残基位置均相对保守。系统进化显示,LvML1蛋白与斑节对虾、日本囊对虾、美洲螯龙虾等甲壳动物的ML蛋白聚为一支。实时荧光定量PCR结果显示,LvML1基因在所有检测组织中均有表达,在心脏中表达量最高。发育表达结果显示,LvML1基因自无节幼体期已有表达,在变态至溞状幼体期、糠虾幼体期及仔虾的每次转换均上调表达,表明其参与了对虾幼体的发育过程。十足目虹彩病毒1人工感染24 h后,LvML1基因在肝胰腺和鳃中表达量下降,差异极显著(P<0.01),说明其参与了对虾抗病毒的先天免疫过程。本试验结果可为了解凡纳滨对虾ML蛋白家族的结构功能及其在对虾先天免疫和幼体发育中的重要作用提供基础资料。ML(MD-2-related lipid-recognition)genes are a class of proteins with important functions in innate immunity and lipid metabolism in invertebrates.In this study,we cloned a novel ML gene from Pacific white shrimp Litopenaeus vannamei named LvML1.The full-length cDNA sequence of LvML1 consists of a 254 bp 5′-untranslated regions(5′-UTR),a 913 bp 3′-UTR and a 462 bp open reading frame(ORF).The deduced amino acid sequence is composed of 153 amino acids.LvML1 has seven cysteine residues,which are predicted to form three pairs of disulfide bonds,with a typical ML family structural domain.Multiple sequence alignment revealed that both the ML structural domain and cysteine residue positions were relatively conserved.Phylogenetic tree showed that LvML1 clustered with ML proteins of crustaceans including tiger shrimp Penaeus monodon,kuruma shrimp Marsupenaeus japonicus and spiny lobster Homarus americanus.Real-time quantitative PCR results showed that LvML1 was expressed in hepatopancreas,eyestalk,intestine,stomach,heart,gill,muscle and blood,with the highest expression level in the heart.Expression in larval developmental stages showed that LvML1 was already expressed from the nauplius of Pacific white shrimp and was upregulated at each metamorphosis transition to zoea,mysis and postlarva,indicating that it is involved in the larval developmental processes of shrimp.Expression of LvML1 in hepatopancreas and gills exposed to artificial injection of Decapod iridescent virus 1(DIV1)for 0 h and 24 h was significantly down-regulated with P<0.01,indicating that it is involved in the regulation of innate immunity against the virus in shrimps.The finding provides basic data for understanding of the structure and function of the ML protein family in Pacific white shrimp and its important role in the innate immunity and larval developmental processes of the shrimp.

关 键 词：凡纳滨对虾 ML基因 组织表达 发育表达 十足目虹彩病毒1 

分 类 号：Q786[生物学—分子生物学]