重组鸡痘病毒rFPV-VP3-F的遗传稳定性及在鸡体内的表达

Genetic stability of recombinant fowlpox virus rFPV-VP3-F and its expression in chicken

为了评价重组禽痘病毒rFPV-VP3-F的遗传稳定性,将rFPV-VP3-F在鸡胚成纤维细胞(CEF)连续培养20代。对第5、10、15和20代病毒的F基因进行扩增及序列分析,未发现F基因发生氨基酸改变;通过间接免疫荧光方法检测各个代次rFPV-VP3-F中F基因特异表达的情况;将rFPV-VP3-F和禽痘病毒FPV017分别接种于35日龄商品鸡,在免疫后第7、14、21、28、35 d采集各组血清,检测中和抗体效价。结果显示,F基因在重组禽痘病毒中可以稳定遗传,可在CEF中表达相应蛋白,可刺激鸡产生相应的抗体。

The recombinant fowlpox virus rFPV VP3-F was propagated in chicken embryo fibroblast （CEF） for 20 generations to evaluate its genetic stability. In the 5,10,15 and 20th generation, the F gene was cloned, sequenced and no amino acid was changed. Subsequently, indirect immunofluorescence assay was applied to identify expression of F gene in different generations. 20 chickens were randomly separated into two groups,and were immunized with rFPV VP3-F and FPV017 ,respectively,and immunity dose was 106PFU each. Sera were collected on day 7,14,21,28 and 35 post immunization. The neutralizing antibody titers were detected through a neutralization test. The results showed that the F gene had good genetic stability in rFPV VP3-F,could be expressed in CEF,and induced corresponding antibodies in chickens.