【摘要】 NIBRG-14是采用"6+2"策略制备的一株H5N1灭活疫苗株,其表面抗原HA和NA基因来自于A/Vietnam/1194/2004(H5N1,VN1194),内部基因来自于A/Puerto Rico/8/34(H1N1,PR8),已有研究表明该疫苗株在鸡胚中的产量不佳.本研究发现,在PR8背景下,VN1194NA基因被包装入重组病毒中的效率仅为正常包装量的38%～68%,因此有一部分重组病毒为不含有NAvRNA的缺陷型病毒粒子.本研究通过在VN1194NA基因完整编码区(CDS)的5′和3′两端嵌合PR8NA基因包装信号序列(vRNA3′末端41bp,5′末端67bp)的方法,使重组病毒中NAvRNA的包装效率得到完全恢复,并且病毒在鸡胚的生长滴度提高了10倍,血凝素HA含量提高了约2·7倍,从而为H5N1流感疫苗株的研制提供了新的思考方向.更多还原

【Abstract】 NIBRG-14 is one of the H5N1 candidate vaccine viruses developed using "6+2" approach with the hemagglutinin(HA) and neuraminidase(NA) genes derived from A/Vietnam/1194/2004(H5N1,VN1194) and the remaining six internal segments from A/Puerto Rico/8/34(H1N1,PR8).However,NIBRG-14 was reported to yield low amounts of HA antigen.This study found that the NA vRNA of VN1194 is poorly packaged(38%~68%) into the recombinant viruses with a backbone of PR8 genes,causing the formation of defective virions without the NA vRNA in viral genome.Using recombinant DNA techniques,we constructed a chimeric NA gene with the coding region of VN1194 NA flanked by the packaging signal sequence(vRNA 3’end 41bp,5’end 67bp) of PR8 NA.The packaging of NA vRNA is completely restored in the recombinant viruses with the chimeric NA gene.Moreover,the recombinant viruses contained the chimeric NA gene replicate better in chicken embroynated eggs than recombinant viruses with wild type NA gene of VN1194,as indicated by a 10-fold increase in virus titer and 2.7-fold increase in HA antigen content.These findings suggest a novel strategy to improve the in ovo growth and increase the available dose of NIBRG-14 in vaccine manufacture.更多还原