摘要
目的研究中国黄热疫苗生产用减毒株与世界卫生组织(WHO)黄热疫苗标准株的基因序列,了解其核苷酸序列变异程度,和与其相关的氨基酸是否发生突变。方法根据基因库(GenBank)中收录的黄热病毒(Yellow Fever Virus,YFV)17D序列设计引物,提取该两株YFV的核酸,逆转录后进行聚合酶链反应(Polymerase Chain Reaction,PCR)扩增病毒各部分基因片段,PCR产物纯化后直接测序,5'和3'端TA克隆后进行测序,然后进行遗传进化分析。经核苷酸序列测定进行序列比对分析。结果经序列测定后,进行两株YFV的序列比对,并与GenBank中收录的其它相关YFV17D疫苗株和野毒株进行序列比对,发现中国黄热疫苗生产株与WHO黄热疫苗标准株间的核苷酸序列并未发生较大变异,核苷酸和氨基酸的同源性分别约为99%和99%。其重要病毒抗原E蛋白也未发生较大突变,但发现E区与毒力密切相关的173位氨基酸发生回复突变,进一步通过系统发生分析说明两者亲缘关系十分接近。结论传代过程中,中国黄热疫苗株和WHO黄热疫苗标准株在基因水平上除个别外未发生较大改变,其基因组具有一定的稳定性。
Objective To compare the molecular characteristics of the Chinese attenuated yellow fever 17D vaccine strain and the WHO reference yellow fever 17D vaccine strain. Methods The primers were designed according to the published nucleotide sequences of YFV 17D strains in GenBank. Total RNA of was extracted by the Trizol and reverse transcripted. The each fragments of the YFV genome were amplified by PCR and sequenced subsequently. The fragments of the 5＇ and 3＇ end of the two strains were cloned into the pGEM T-easy vector and then sequenced. Results The nucleotide acid and amino acid sequences of the homology to both strains were 99% with each other. No obvious nulceotide changes were found in the sequences of the entire genome of each 17D strains. Moreover, there was no obvious changes in the E protein genes. But the E173 of YF17D Tiantan, associted with the virulence, had mutantions. And the two live attenuated yellow fever 17D vaccine strains fell to the same lineage by the phylogenetic analysis. Conclusion The results indicated that the two attenuated yellow feverl 7D vaccine viruses accumulates mutations at a very low frequency and the genomes were relative stable.
出处
《中国疫苗和免疫》
CAS
2009年第2期145-151,共7页
Chinese Journal of Vaccines and Immunization
关键词
黄热病毒
核苷酸全序列
突变
Yellow fever virus
Completed sequence ofnucleotide acids
Mutation
