黄热病病毒疫苗株遗传稳定性研究

发布时间：2018-04-17 03:15

本文选题：黄热病病毒 + 黄热疫苗　；参考：《北京生物制品研究所》2007年硕士论文

【摘要】： 黄热疫苗是高度减毒的活病毒疫苗，其生产基质是7日龄全鸡胚，在世界范围内已经安全使用了50多年。本实验将17D株黄热疫苗病毒在鸡胚成纤维细胞中连续传代，通过对传代病毒的遗传稳定性研究，探讨将传统用于黄热疫苗生产的全鸡胚改换为鸡胚成纤维细胞后，是否会对疫苗的遗传稳定性产生影响，从而为使用鸡胚成纤维细胞作为黄热疫苗生产基质的科研路线提供一些依据。在本实验中选择黄热疫苗国际标准品99／616(NCBI编号)和我国黄热疫苗工作毒种2006(54)0239(天坛生物编号)作为实验用传代毒种，两者都是17D疫苗株，其中99／616分离自17D-204株。将两株毒种分别在鸡胚成纤维细胞中适应性传代，各自传10代，记作“99／616-CEF传代病毒”和“YFVBJ-CEF传代病毒”。选取99／616-CEF传代病毒和YFVBJ-CEF传代病毒的1、3、6、9代病毒，对其结构基因片断C、M、E的核苷酸序列进行测定。此外，对我国商品用黄热疫苗的核苷酸序列也进行测定，该疫苗的工作毒种是2006(54)0239，培养基质为7日龄活鸡胚。对测序结果的数据分析包括三部分，一：分析99／616-CEF传代病毒的遗传稳定性。将99／616-EEF传代病毒的测序结果与NCBI GENEBANK公布的17D-204株黄热疫苗的核苷酸序列进行比较。二：分析YFVBJ-CEF传代病毒的遗传稳定性。将YFVBJ-CEF传代病毒与我国商品用黄热疫苗的测序结果进行比较。三：分析我国黄热疫苗毒种的历史背景。将我国商品用黄热疫苗的测序结果与NCBI GENEBANK公布的17D-204、17DD株黄热疫苗的核苷酸序列分别进行比较。通过实验结果分析推断：黄热疫苗病毒经过在鸡胚成纤维细胞中传代后，，其遗传物质会发生适应性改变。99／616-CEF传代病毒和YFVBJ-CEF传代病毒在结构基因片断C上均有较活跃的核苷酸位点；结构基因片断M、E在99／616-CEF传代病毒中表现比较活跃，而在BJYFV-CEF传代病毒中没有发生改变。我国的黄热疫苗毒种来自于黄热病毒减毒疫苗株17D株，在基因水平上与17D-204、17DD株有不同之处，可能是17D-204疫苗株的另一旁支。
[Abstract]:Yellow fever vaccine is a highly attenuated live virus vaccine. Its production substrate is 7 days old whole chicken embryo. It has been used safely in the world for more than 50 years.In this experiment, the 17D yellow fever vaccine virus was continuously subcultured in chicken embryo fibroblasts. By studying the genetic stability of the virus, the whole chicken embryo used in yellow fever vaccine production was changed into chicken embryo fibroblast.Whether it will affect the genetic stability of the vaccine will provide some basis for the research route of using chicken embryo fibroblasts as the substrate for producing yellow fever vaccine.In this experiment, yellow fever vaccine international standard 99/616(NCBI number) and Chinese yellow fever vaccine working strain 2006 ~ 54,0239 (Temple of Heaven biological code) were selected as experimental subculture virus. Both of them were 17D vaccine strains, of which 99 / 616 were isolated from 17D-204 strain.The two strains were subcultured adaptively in chicken embryo fibroblasts for 10 generations, respectively, and were recorded as "99/616-CEF subculture virus" and "YFVBJ-CEF subculture virus".The nucleotide sequence of the structural gene fragment of 99/616-CEF virus and YFVBJ-CEF virus was determined.In addition, the nucleotide sequence of yellow fever vaccine for commercial use in China was also determined. The working strain of the vaccine was 2006A54,0239, and the culture medium was 7-day-old live chicken embryo.The data analysis of sequencing results consists of three parts: first, the genetic stability of 99/616-CEF subculture virus is analyzed.The sequencing results of 99/616-EEF subculture virus were compared with the nucleotide sequence of yellow fever vaccine of 17D-204 strain published by NCBI GENEBANK.Second, the genetic stability of YFVBJ-CEF subculture virus was analyzed.The sequencing results of YFVBJ-CEF subculture virus and commercial yellow fever vaccine in China were compared.The third is to analyze the historical background of yellow fever vaccine in China.The sequencing results of commercial yellow fever vaccine in China were compared with the nucleotide sequence of 17D-204DD-17DD strain yellow fever vaccine published by NCBI GENEBANK.The results showed that the genetic material of yellow fever vaccine virus could change adaptively after it was subcultured in chicken embryo fibroblasts. Both of them had active nucleotide sites on structural gene fragment C.The structural gene fragment MNE was active in the 99/616-CEF passaged virus, but not changed in the BJYFV-CEF subculture virus.The yellow fever vaccine strains in China come from the attenuated yellow fever virus strain 17D, which is different from 17D-204A17DD strain in gene level and may be another branch of 17D-204 vaccine strain.
【学位授予单位】：北京生物制品研究所
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R392

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