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目的对我国现用风疹病毒(Rubella virus,RV)疫苗生产毒株松叶株(Matsuba)进行全基因组测序,分析其在减毒过程中的遗传与变异特点,为其安全性评价及遗传学质量控制的标准化提供依据。方法利用RT-PCR法分段扩增我国现用Matsuba疫苗株主代种子基因组全序列,分别将产物插入到T-A克隆载体pGEM-T easy中,构建病毒cDNA文库,进行全基因组序列测定与分析。结果我国现用Matsuba疫苗株主代种子基因组全长9 762 nt,含2个ORF,分别位于核苷酸41～6388位和6512～9700位,编码2 116、1 062个氨基酸;与GenBank中登录的Matsuba.GMK3野毒株和Matsuba疫苗株核苷酸同源性分别为97.5%和99.9%,氨基酸同源性分别为98.8%和99.9%;其在减毒的过程中共有37个氨基酸位点发生变异,其中非结构蛋白P150有18个氨基酸位点发生突变,结构蛋白E1和E2主要抗原位点氨基酸未发生变异,E1-177位糖基化位点突变丢失。结论我国现用风疹病毒Matsuba疫苗株与同类减毒株具有较高的同源性,其主要的抗原位点在减毒保种过程中高度保守,为在分子水平上保证Matsuba株毒种及其生产疫苗的安全性提供了依据。
Objective To analyze the genetic and variation characteristics of Matsuba, an active vaccine strain of Rubella virus (RV) in China, and to evaluate its safety and genetic quality Standardization of control provided. Methods The full-length genomic sequence of the main Matsuba vaccine strain in China was amplified by RT-PCR. The product was inserted into the T-A cloning vector pGEM-T easy to construct a virus cDNA library for genome-wide sequence analysis and analysis. Results The full-length genome of Matsuba vaccine was 9762 nt in length with 2 ORFs located at nucleotide positions of 41 to 6388 and 6512 to 9700, encoding 2,116 and 1,062 amino acids, respectively. GenBank accession number The nucleotide homology of Matsuba, GMK3 and Matsuba vaccine strains were 97.5% and 99.9%, respectively, and their amino acid homologies were 98.8% and 99.9% respectively. A total of 37 amino acid sites were found during attenuating Among them, the non-structural protein P150 has a mutation of 18 amino acid residues, the amino acids of the major antigenic sites of the structural proteins E1 and E2 are not mutated, and the mutation of the E1-177 glycosylation site is lost. Conclusions The Matsuba vaccine strain currently used in our country has high homology with the attenuated strains of the same type. The major antigenic sites are highly conserved during the process of attenuated seed protection. In order to ensure the Matsuba strain at the molecular level and its Provide a basis for the safety of vaccine production.