犬细小病毒VP2基因的克隆与生物信息学分析  

Cloning and Bioinformatics Analysis of Canine parvovirus VP2 gene

作　　者：叶新慧[1] 申魁魁[1] 符欣蕙[1] 王丽霞[1] 李恭贺[1] 张明[1] 卢晟盛[1,2] 卢克焕[1,2] 郑喜邦[1] 

机构地区：[1]广西大学动物科技学院,南宁530004 [2]亚热带农业生物资源保护与利用国家重点实验室,南宁530004

出　　处：《基因组学与应用生物学》2012年第6期554-558,共5页Genomics and Applied Biology

基　　金：广西自然科学基金项目(2011GXNSFD018018);亚热带农业生物资源保护与利用国家重点实验室开放课题(SB0907)共同资助

摘　　要：本研究旨在克隆犬细小病毒VP2(Canine parvovirus VP2,CPV-VP2)基因,构建克隆载体pMD18-T-VP2,分析其生物信息学特征。以犬细小病毒(CPV)阳性病犬血液基因组病毒DNA为模板,PCR扩增CPV-VP2基因,通过TA克隆获得pMD18-T-VP2重组质粒,测序后对其进行生物信息学分析。结果表明:(1)CPV-VP2完整的开放阅读框由1755个核苷酸组成,共编码585个氨基酸残基,其序列同源性与浣熊最高,达97.6%;(2)信号肽在线预测认为,CPV-VP2基因无信号肽;(3)CPV-VP2大多数氨基酸偏好以T、A结尾的密码子;(4)CPV-VP2蛋白可能存在7个糖基化位点,其丝氨酸、苏氨酸、酪氨酸分别存在8个、10个和7个磷酸化位点;(5)CPV-VP2可能存在76个B细胞抗原表位;(6)CPV-VP2蛋白为亲水性蛋白,无跨膜结构;(7)CPV-VP2蛋白二级结构的α-螺旋区域占9.93%、β-折叠区域占24.49%、无规则卷曲区域占65.58%;(8)CPV-VP2蛋白三级结构存在多个螺旋和折叠区域,主要以无规则卷曲为主。本研究为制定CPV-VP2基因原核或真核高效表达策略提供了参考资料,并为进一步制备CPV胶体金快速诊断试纸条和基因工程疫苗奠定了基础。The present study was to clone VP2 gene of Canine parvovirus （CPV）, to construct cloning vector pM-D18-T-VP2, and finally to perform a bioinformatical analysis after sequencing. A blood sample was collected from a dog infected CPV disease, and virus DNA was used to amplify CPV-VP2 gene by PCR. Constructed by TA cloning, the recombinant plasmid pMD18-T-VP2 was sequenced and followed by a bioinformatical analysis. The results showed that： （1） The whole open reading frame of CPV-VP2 gene is composed of 1 755 nucleotide acids, en-coding 585 amino acids, which shares the highest sequence similarity （97.6%） in nucleotide and amino acids with the VP2 homologue of Procyon Lotor; （2） The online signal peptide prediction showed that there is no signal pep-tide in CPV-VP2 gene; （3） Most amino acids of CPV-VP2 gene prefer to codons with end of T or A; （4） 7 glyco-sylation sites may exist in CPV-VP2 gene, with 8, 10 and 7 phosphorylation sites respectively in serine, threonine and tyrosine; （5） CPV-VP2 gene possibly contains a total of 74 B cell epitopes; （6） CPV-VP2 protein is hydrophilic protein without transmembrane domains. （7） 9.93% of α-helixes, 24.49% of β-sheets and 65.58% of radon coils construct the secondary structure of CPV-VP2 protein; （8） The tertiary structure of VP2 protein contains some helixes and sheets, while random coils is dominated. This study would lead to preparation of CPV monoclonal antibody and colloidal gold diagnostic test strip.

关 键 词：犬细小病毒 VP2基因 分子克隆 生物信息学分析 

分 类 号：S852.65[农业科学—基础兽医学]