RESUMEN
Twenty Newcastle disease virus(NDV)strains were isolated from diseased chicken and geese in field outbreaks during 2005 and 2006 in some regions of Jiangsu and Guangxi,and the antigenic analysis of the all NDV isolates had been done based on the reaction spectrum with a panel of monoclonal antibodies to the HN glycoprotein.The entire ORFs encoding HN protein of these NDV isolates were amplified by RT-PCR successfully,cloned and sequenced.The resultant sequences of HN genes of 13 isolates of chicken origin and 7 isolates of goose origin were gained and analyzed.The results of reaction spectrum showed that there were some distinct differences in the antigenic epitopes among the 20 NDV isolates.And the sequences revealed that the coding regions of the HN genes of these isolates all consisted of 1716 nt characteristic of virulent strains of NDV,coding for 571 amino acids.Neucleotides sequence homology were found to be from 94.8%to 100%among 18 NDV isolates of genotypeⅦ,and the neucleotides sequence homology between all the isolates and the other genotypeⅦstrains of recent years in China ranged from 92.1%to 99.6%.The deduced amino acid sequences and the receptor-binding regions of HN proteins between the NDV isolates of chicken origin and of goose origin were compared and analyzed.The results showed that some unique amino acid substitutions were found in the genome of the NDV isolates,and the close genetic similarity provided evidence for epidemiological linkage between the NDV isolates of chicken origin and of goose origin in the same period.
RESUMEN
Objective To construct an eukaryotic expression plasmid containing gene coding for the hemagglutinin-neuraminidase(HN)of newcastle disease virus(NDV)oncolytic strain Italien,and then to express the protein in eukaryotic cell.Method The HN cDNA was synthesized from viral RNA by RT-PCR,and the eukaryotic expression vector of HN gene(named pcDNA3.1-HN)was constructed.The vector pcDNA3.1-HN was transfected into CHO-K1 cell by liposome,and G418 was used to select stable clones expressing HN gene.The expression of HN protein was visualized by Western blot and Immunofluorescence microscopy.Results Restriction analysis and DNA sequencing proved that HN gene was correctly cloned into expression vector.Western blot analysis and immunofluorescence showed that the HN was expressed in CHO-K1 cells.Conclusion The HN cDNA of NDV was successfully cloned into eukaryotic vector which showed good expression of HN protein in CHO-K1 cells.