禽冠状病毒分离鉴定及相关结构基因遗传变异分析

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1、上海交通大学硕士学位论文禽冠状病毒分离鉴定及相关结构基因遗传变异分析姓名：钱华冬申请学位级别：硕士专业：农业与生物学指导教师：华修国;朱建国20060101上海交通大学硕士学位论文因S开放阅读框ORF由3504个核苷酸组成编码1167个氨基酸残基组成的多肽共有31个潜在的N-糖基化位点与二硫键形成有关的半胱氨酸残基数量为35个M基因全序列共有681bp的核苷酸片段其中ATGC所占百分比分别是28.65%22.32%18.21%30.84%G+C含量为40.53%,共编码226个氨基酸有2个潜在的N-端糖基化与二硫键形成有关的半胱氨酸残基数量为9个N基因经测序得到大小为1275bp的

2、核苷酸片段,包含了核蛋白基因完整阅读框,其中编码区为1230个核苷酸,编码409个氨基酸,其中赖氨酸(Lys)占所有氨基酸残基的9.54%(39/409);丝氨酸(Ser)占8.31%(34/409);精氨酸(Arg)占7.82%(32/409),远高于其他氨基酸残基含量将PSH主要结构基因序列与GenBank中公开发表的其它冠状病毒代表株相应序列同源性比较后发现PSH与第3抗原群冠状病毒有高度同源性其中与IBV的亲缘关系较TCV更近而与包括SARS在内的其它冠状病毒主要结构基因同源性较低关键词:禽;冠状病毒结构基因遗传变异上海交通大学硕士学位论文ISOLATION,IDENTIF

3、ICATIONANDGENETICVARIATIONOFRELATEDSTRUCTURALGENEOFAVAINCORONAVIRUSESABSTRACTInordertoinvestigatethegeneticvariationandepidemictrendofaviancoronavirusesinShanghai,andprovidewiththeoreticsbasisforthepreventionandcureofthedisease,thisresearchwasundertakenincludingthefollowing:(1)Forthepurposeofv

4、irusisolation,tissuesamplesandlaryngotrachealscrapingsofpoultrywhichcollectedinShanghaiwereinoculatedintoSPFchickenembryosaftergeneralprocess.Threeblindpassagesweredone.Fivecoronavirusesisolateswereidentifiedfromthediseasedfowlsinducedbyspecificpathogen-freeembryonatedchickeneggsinoculation,th

5、roughelectronmicroscopicobservationandRT-nested-PCRmethod,haemagglutinationandpathogenicitytests.Thefourcoronavirusisolates(avianbronchitisvirus,IBV),designatedasSH1SH2SH3andSH4respectively,wereisolatedfromthechicken.TheotherwasisolatedfromthepigeonsnamedPSH.(2)AccordingtothepublishedS1sequenc

6、esofavianbronchitisvirusesinGenbank,apairofspecificprimerswasdesignedtoamplifyS1geneoftheIBVisolatesbyRT-PCRmethod.ThenthecDNAofS1geneswerepurifiedandclonedintopMD18-Tvectorandsequencedrespectively.SequencesdatashowedthatS1geneofIBVSH1strainwascomposedof1635nucleotidescoding544aminoacid;S1gene

7、ofIBVSH2strainwascomposedof1626nucleotidescoding541aminoacid;S1geneofIBVSH3strainwascomposedof1614nucleotidescoding537aminoacid;S1geneofIBVSH4strainwascomposedof1626上海交通大学硕士学位论文nucleotidescoding541aminoacid;TheIBVstrainsshared77.4%82.9%