| Newcastle disease (ND),otherwise, Asian Fowl pest , caused by Newcastle disease virus(NDV), can cause a acute, highly contagious and fatal infectious of avian diseases. It spread widely in the world and got vast lose in poultry of our country. The molecular epidemiology research of ND, that not only provides guidance for the prevention and control of ND, but also can reveal the heredity and evolution of NDV in our country, is one of important path to research the epidemiology of ND. Researching the function of the P gene of NDV not only in favour of revealing the dynamic reaction between NDV and organism, but also underlaying the development of new vaccine . In this study, three strains NDV animal was proliferated, the P gene of three strains NDV was cloned and the variation of P gene was analyzed. Templating the P gene, the V gene of three strains of NDV was cloned by means of artificial rite-directed mutagenesis, and the character of primary structure and secondary structure of V protein was analyzed by bioinformatics tools . Eukaryotic expression plasmid of V gene were designed in basis of PEGFP-C1; the eukaryotic expression requirements were studied and the expression product were identified. The main study as follows:1,Three strains of NDV were proliferated (NnShX05,ChickenShX08,PigeonShX03), and total length of P gene were cloned by means of TA clone . The sequences of P gene of three NDV were compared with other P gene from GeneBank, and phylogenetic tree was drew. The result showed that , homologous rate of the P gene of NnShX05 and referred strains nucleotides and deduced amino acids sequences was 82.1%～99.8% and 81.1%～100% respectively; homologous rate of the P gene of ChickenShX08 and referred strains nucleotides and deduced amino acids sequences was 82.1%～100% and 81.1%～100% respectively ; homologous rate of the P gene of PigeonShX03 and referred strains nucleotides and deduced amino acids sequences was 82.4%～93.1% and 79.1%～93.2% respectively ; There were two nucleotide variation between NnShX05 and ChickenShX08, and there were evident variation between NnShX05,ChickenShX08 and PigeonShX03 . NnShX05 and ChickenShX08 were highly homologous with HB92 isolate V4 but bare homologous with EU296519.1TW06-316; PigeonShX03 was highly homologous with EU296531.1TW84-P72 but bare homologous with EU4199882.1D58. The result of phylogenetic analysis indicated that NnShX05 and ChickenShX08 would evolve from vaccine strains, PigeonShX03 would evolve from strains of Taiwan or they would share the same progenitor.2,Templating the P gene , the V gene of three strains of NDV were cloned by means of big prime artificial rite-directed mutagenesis . The result of comparing them with each other showed: the nucleotide there was only one nucleotide diversity between NnShX05 and ChickenShX08, the deduced amino acids sequences of NnShX05 and ChickenShX08 were the same with each other, they were much diversity between them and PigeonShX03. The predicted construction of V protein indicated that : There were the same number of amino acid between NnShX05 and PigeonShX03 , the glycosylation site and Zn finger domain lied in the same region , but there were much diversity between them in amino acid composition,isoelectric point,acylations site (one site variation)and phosphorylation site(seven sites variation) .The result indicated that the ability of V protein from different animal antagonism to interferon may be difference.3,In basis of PEGFP-C1 , the eukaryotic expression vectors were designed with green fluorescent protein as reporter genes . By transfecting eukaryotic cells, the eukaryotic expression requirements were studied in vitro. The result showed that V gene could express in Vero cell favourably. Identified by western-blotting, it showed that the expressed protein were with reactionogenicit, and could react with antiserum specific to NDV. The study provide basis for researching difference of V protein from different animal antagonism to interferon.
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