摘要
分别对鸡源、鸭源、鹅源新城疫病毒（NDV）强毒分离株（WF00C、WF00D、WF00G）的F基因进行RT-PCR扩增、克隆和序列测定,并对其核苷酸和推导氨基酸序列进行分析。结果表明,各分离株F基因均含1个完整的开放阅读框（ORF）,全长为1 662 bp,编码553个氨基酸,有6个潜在的糖基化位点,裂解位点区的氨基酸序列均为112R-R-Q-K-R-F117,与强毒株特征相符,具有第101位的K（赖氨酸）和121位的V（缬氨酸）,符合基因Ⅶ型NDV特征;WF00C株有13个半胱氨酸（Cys）残基,WF00D株和WF00G株各有12个Cys残基;在限制性内切酶（RE）位点（nt334~1 682）的分布上,3个毒株均存在多数鹅源分离株特有的973 nt和1 249 nt RsaⅠ位点;WF00D株与WF00C株、WF00G株以及GenBank的15株各基因型代表株的F基因编码区同源性进行比较,其核苷酸序列同源性为84.7%~99.3%,推导的氨基酸序列同源性为88.4%~98.9%;3个分离株间的同源性较高,其与ZJ1、SF02和NA_1等鹅源毒株之间差异较小,而与LaSota、F48E9等毒株之间差异明显。根据NDV系统发育进化树、酶切位点分析以及基因分型结果,证明它们的基因型均为Ⅶd亚型。
The whole sequence of F gene of avian paramyxovirus isolates was amplified by reverse transcription-polymerase chain reaction（RT-PCR）,and further cloned into pMD18-T vector,and positive recombinant plasmids were obtained by polymerase chain reaction（PCR）.The sequence analysis showed that all the nucleotide sequence of the F gene were 1 662 bp in length and had a single opening read frame（ORF）,which encoded polypeptide of 533 animo acid,contained 6 potential asparagine-linked glycosylation sites,their amino acid sequence of cleavage site region residues were 112R-R-Q-K-R-F117,matched to the characteristics of virulent strain,the isolates possessed K101and V121,which were similar to the characteristic of genotype Ⅶ;WF00C strain contained 13 cysteine（Cys） residues,but WF00D and WF00G strains only had 12Cys residues,all isolates contained 973 nt and 1 249 nt RsaⅠrestriction endonuclease（RE）sites（nt334-1 682）.Comparison of F gene nucleotide sequences of WF00D strain with 15 other published sequences from GenBank showed that the homology of the nucleotide was 85.0%-99.3%,the homology of the deduced amino acid sequences was 88.8%-98.9%.Three isolates from Anhui shared higher homonology each other,and higher homonology compared with ZJ1、SF02、NA_1 too,but lower homonology compared with F48E9、LaSota.The results of phylogenetic tree,restriction site analysis and genotype characterization showed that three strains WF00C,WF00D,WF00G belonged to novel subgenotype Ⅶd.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2010年第8期1043-1048,共6页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(30671556)
关键词
禽副黏病毒
新城疫病毒
F基因
克隆
序列分析
avian paramyxovirus
newcastle disease virus
F protein gene
cloning
sequencing
