家蚕核型多角体病毒lef-8基因克隆及系统进化分析  

Cloning and phylogenetic analysis of lef-8 gene in Bombyx mori nucleopolyhedrovirus

作　　者：张永红 杨大平 许珍娣 邵榆岚 李玲利 张宏瑞[1] ZHANG Yong-hong;YANG Da-ping;XU Zhen-di;SHAO Yu-lan;LI Ling-li;ZHANG Hong-rui(College of Plant Protection,Yunnan Agricultural University,Kunming,Yunnan 650201,China;Institute of Sericulture and Apiculture,Yunnan Academy of Agricultural Science,Mengzi,Yunnan 661101,China;Longchuan Agricultural Technology Extension Center,Longchuan,Yunnan 678700,China)

机构地区：[1]云南农业大学植物保护学院,云南昆明650201 [2]云南省农业学院蚕桑蜜蜂研究所,云南蒙自661101 [3]陇川县农业技术推广中心,云南陇川678700

出　　处：《南方农业学报》2023年第7期2126-2134,共9页Journal of Southern Agriculture

基　　金：国家自然科学基金项目(31960684);微孢子虫感染与防控重庆市重点实验室开放课题基金项目(CKMIC202102)。

摘　　要：【目的】了解云南省德宏州陇川县蚕区家蚕核型多角体病毒(BmNPV)株系的遗传地位和多样性,为厘清云南蚕区BmNPV的起源及科学防控家蚕血液型脓病提供参考依据。【方法】对分离自云南省德宏州陇川县蚕区的BmNPV进行lef-8基因克隆,通过ExPASy、SignalP 5.0、TMHMM 2.0、NetOGlyc 4.0、NetPhos 2.0及SWISS-MODEL等在线软件进行生物信息学分析,并基于lef-8基因核苷酸序列相似性,采用MEGA7.0中的邻接法(NJ)构建系统发育进化树。【结果】云南省德宏州陇川县蚕区BmNPV-YNLC株的lef-8基因序列全长2631 bp,共编码876个氨基酸残基;其编码蛋白(LEF-8)无信号肽序列,也不存在跨膜结构域,蛋白分子量为101.63 kD,理论等电点(pI)为8.8;在第750~800位氨基酸残基区域具有较高的疏水性,部分氨基酸位点表现为亲水性,且具有较高抗原指数和表面可及性;存在6个潜在的N-糖基化位点,62个磷酸化位点(25个丝氨酸磷酸化位点,23个苏氨酸酸化位点,14个酪氨酸磷酸化位点),但不存在O-糖基化位点。BmNPV-YNLC株与BmNPV-T3株的lef-8基因核苷酸序列相似性为99.7%,BmNPV-YNLC株lef-8基因在第96~98位核苷酸序列上存在3个碱基(CAA)缺失;此外,还存在2个非同义突变(^(646)T→A和^(1895)C→G)和7个同义突变。基于lef-8基因核苷酸序列相似性构建的系统发育进化树显示,BmNPV-YNLC株与日本和韩国地区的BmNPV亲缘关系较近,而与我国流行的BmNPV遗传关系较远。【结论】lef-8基因在不同地区来源的BmNPV株系中高度保守,BmNPV-YNLC株与日本和韩国地区的BmNPV亲缘关系较近,但其LEF-8蛋白N端存在氨基酸缺失突变,说明BmNPVYNLC株为云南蚕区的新流行株系。【Objective】This study aimed to understand the genetic diversity of the Bombyx mori nucleopolyhedrovirus(BmNPV)strains in sericultural regions of Longchuan County,Dehong Prefecture,Yunnan Province,and provide reference for the origin of Yunnan BmNPV and the scientific prevention and control of B.mori nuclear polyhedrosis.【Method】The lef-8 gene of BmNPV isolate from B.mori in the sericultural regions of Longchuan County,Dehong Prefecture,Yunnan Province was cloned and analyzed based on bioinformatics through online software including ExPASy,SignalP 5.0,TMHMM 2.0,NetOGlyc,NetPhos 2.0 and SWISS-MODEL.Then neighbor-joining(NJ)method of MEGA 7.0 was applied to constructa phylogenetic treeon the basis of the similarity of lef-8 gene nucleotide sequence.【Result】The whole length of lef-8 gene nucleotide sequence of BmNPV-YNLC strain from sericultural regionsof Longchuan County,Dehong Prefecture,Yunnan Province was 2631 bp,which encoded 876 amino acid residues.The encoded protein(LEF-8)had neither signal peptide sequence nor transmembrane domain,with the molecular weight of the protein being 101.63 kD and the theoretical isoelectric point(pI)8.8.The amino acid residues from 750 to 800 showed high hydrophobicity,and some amino acid sites showed hydrophilicity,high antigenic index and surface accessibility.The amino acid sequence of the protein contained 6 potential N-glycosylation sites and 62 phosphorylation sites(25 serine phosphorylation sites,23 threonine phosphorylation sites and 14 tyrosine phosphorylation sites),but did not have O-glycosylation sites.The similarity of lef-8 gene nucleotide sequence from BmNPV-YNLC strain and BmNPV-T3 strain was 99.7%.The lef-8 gene of BmNPVYNLC strain had three base(CAA)deletionat positions 96-98,and there were two nonsynonymous mutations(^(646)T→A and^(1895)C→G)and seven synonymous mutations in the nucleotide sequence.The phylogenetic tree constructed via lef-8 gene nucleotide sequence similarity suggested that the genetic relationship between BmNPV-YNLC strain and B

关 键 词：家蚕 家蚕核型多角体病毒(BmNPV) lef-8基因 遗传多样性 系统进化分析 

分 类 号：S884.51[农业科学—特种经济动物饲养]