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目的研究沪191(S191)麻疹病毒疫苗株的核蛋白(N)基因。方法将S191株主代病毒(HK33HAM39CEC20)在实验室连续传递11代,从原代鸡胚细胞(CEC)20代连续传至CEC31代。除观察每代病毒培养特性外,还选其中的21、22、25、26、29、30、31代病毒及S191强毒株(HK33HAM45)、Edmonston(Ed)强毒株(HK28HAM40MRC-55),以逆转录-聚合酶链反应(RT-PCR)技术对这些病毒标本的N基因编码区的全长以及羧基末端的450个核苷酸区域进行片段扩增。结果通过N基因的序列分析发现,S191疫苗株与强毒株在N基因上有10~13个核苷酸差异,差异率为0·63%~0·82%。S191株主代病毒在CEC上连续传递11代的过程中,有2个代次病毒的个别核苷酸出现有义突变,分别导致第22代病毒的278位氨基酸Glu→Lys,第26代病毒的341位氨基酸Val→Lys,但这一变化不存在重复性,随着继续传代又恢复到21代的序列水平。结论S191麻疹病毒疫苗株在CEC长期传代过程中,各代次之间的N基因高变区序列一致,包括N基因中与免疫功能有关的3个B细胞表位区域的序列在传代中也无改变,故不可能影响到疫苗的免疫效果。
Objective To study the nuclear protein (N) gene of Shanghai 191 (S191) measles virus vaccine strain. Methods The S191 strain of host virus (HK33HAM39CEC20) was serially delivered in the laboratory for eleven passages and was serially passaged from the passage 20 of the primary chick embryo cells (CEC) to the passage of CEC31. In addition to observing the characteristics of each generation of virus culture, selected 21,22,25,26,29,30,31 generation of virus and S191 virulent strain (HK33HAM45), Edmonston (Ed) virulent strain (HK28HAM40MRC-55), The full length of the N gene coding region and the 450 nucleotide region of the carboxyl terminal of these virus samples were amplified by reverse transcription-polymerase chain reaction (RT-PCR). Results The sequence analysis of N gene showed that the S191 vaccine strain and the virulent strain had 10 ~ 13 nucleotide differences in the N gene, with a difference of 0.63% ~ 0.82%. S191 strain of the main virus in the CEC on the continuous transmission of 11 generations of the process, there are two generations of virus-specific nucleotide mutations appear, leading to the 22nd generation of the virus 278 amino acids Glu → Lys, 26th generation of virus The amino acid Val → Lys at position 341, however, did not show any reproducibility and returned to the 21-generation sequence as the passage continued. CONCLUSIONS: During the long-term passage of CEC in S191 measles virus vaccine, the sequence of hypervariable region of N gene in all generations is the same, including the sequence of 3 B-cell epitopes related to immune function in N gene. Change, it is impossible to affect the vaccine’s immune effect.