Cloning of neuraminidase (NA) gene and identification of its antiviral activity
Neuraminidase not only works as an antigen, inducing target-specific antibodies, but also plays a role of enzyme activity and destroys the sialic acid receptor required by virus infection of the host cell surface which protects the host from virus damage. In order to explore a new idea to use neuraminidase (NA) gene and produce disease-resistant transgenic poultry, prokaryotic expression vector pGEX-NA was constructed to make NA polyclone antibody. Eukaryotic expression vector pcDNA3.0-NA and pcDNA3.0/EGFP-NA was constructed to reveal its subcelluar location by immunofluorescence and enhanced green fluorescent fusion protein (EGFP). Chicken embryonic fibroblast (CEF) cells were transfected with pcDNA3.0-NA and selected by G418 for two weeks, the transfected cells were challenged by Newcastle disease virus (NDV), the morphology of CEF cells were observed to detect the antiviral ability of NA gene. CEF cells were incubated by the cell lysates extracted from the NIH 3T3 cells, which were transfected with pcDNA3.0-NA. The results show that pGEX-NA could express NA protein in vitro and NA polyclone antibody worked very well; immunofluorescence and EGFP fusion protein revealed that NA protein located at the cytoplasm near the membrane; NDV-CEF inhibition experiment showed the NA protein could resist and delayed CEF cells from NDV infection.
Key words: Neuraminidase (NA), newcastle disease virus (NDV), antiviral activity, chicken embryonic fibroblast (CEF).