Cloning and Expression of Neuraminidase Gene of H5N1Subtype Avian Influenza Virus
Abstract: A pair of clone primers were designed and synthesized based on neuraminidase(NA) gene sequences of known Avian influenza virus H5N1 subtypes. NA gene were amplified from total RNA, which had been extracted from allantoic fluid of H5N1 subtype virus inoculated embryo, by reverse transcriptase-polymerase chain reaction using high proofreading polymerase (PyobestTM DNA Polymerase), and expressed using Invitrogen championTM pET directional TOPO expression system. 53.8kDa recombinant NA fusion with polyhistidine (6xHis) tag in N-terminal was expressed and purified. Its immunoreactivity and immunogenicity had been analyzed. The results showed the recombinant NA could not only bind to antiserum against H5N1 subtype virus with specificity but also elicit specific antibody in immunizing animal, and possessed good antigenicity.