甲型H1N1流感病毒NA、PB2和PA表位抗原区段的克隆和表达

Cloning and Expression of NA,PB2 and PA of Novel H1N1 Influenza A Virus

目的:分析比对甲型H1N1流感病毒神经氨酸酶(NA)、聚合酶B2(PB2)和聚合酶A(PA)抗原的序列,克隆表达保守的和变异的表位抗原区段,为免疫学诊断试剂的研究提供候选抗原。方法:采用BioSun生物学软件比较新近公布的A/H1N1流感病毒和我国猪流感病毒浙江株NA、PB2和PA抗原序列,并预测筛选NA、PB2和PA抗原表位保守区段与变异区段,采用PCR逐步合成法合成NA、PB2和PA表位抗原区段基因序列,并利用原核表达载体pBVIL1进行克隆表达。结果:筛选的保守表位抗原区段和变异表位抗原区段为NA/135～180aa、NA/310～350aa、PB2/1～80aa、PA/41～90aa、PA/231～280aa和PA/341～400aa;获得6条表位抗原区段基因序列,且6条表位抗原区段均获得了高效表达,得到纯化后的抗原。结论:获得了A/H1N1流感病毒NA、PB2和PA表位抗原区段,为进一步研制特异的甲型流感病毒快速诊断试剂提供了抗原储备。

Objective：In order to provide the candidate antigens for immunological diagnosis,to analysis the sequence of neuraminidase（NA）,polymerase B2（PB2）and polymerase A（PA）of H1N1 influenza A virus,and to clone and express the conservative and variable antigen epitope segments of the NA,PB2 and PA.Methods：The amino acid sequences of the newly published NA,PB2 and PA,and Zhejiang swine influenza virus NA,PB2 and PA were analyzed and aligned using the BioSun software.The genes of antigen epitope segments were synthesized by the stepwise synthesis PCR.The vector pBVIL1 was used to clone and express the antigen epitope segments.Re- sults：The antigen epitope segments of NA/135～180 aa,NA/310～350 aa,PB2/1～80 aa,PA/41～90 aa,PA/231～ 280 aa and PA/341～400 aa were selected.The genes of the six segments were synthesized,and all of the segments were expressed and purified highly.Conclusion：The antigen epitope segments of NA,PB2 and PA were obtained and have potential application in the development of H1N1 influenza A virus diagnostic reagent.