Cloning of fusion glycoprotein gene of avian paramyxovirus NA--1 strain and contructed of re--bacmid

The geese′s paramyxovirus isolate NA-1 was propagated in 11-day-old chicken embryos.The allantonic fluid of the virus were concentrated and the genomic RNA was extracted.The F gene of the virus has been successfully amplified by RT-PCR,and further cloned into pMD18-T vector,and a positive recombitant plasmid was screened by restriction enzyme analysis.The sequence analysis showed that the nucleotide sequence of this F gene was 1 672 bp and encoding a protein of 553 amino acids.The amino acid sequence of cleavage site region was ~(112)R-R-Q-K-R-F~(117),matching to virulent NDV strains,and at the same time it was consistent with the pathogenicity test isolates.The geese′s paramyxovirus isolate NA-1 belongs to genotype Ⅶ.A recombinant transposon vector(pFF)was contructed by inserting the F gene of strain siping into the transposon vector pFastBacⅠ.Then,pFF was transformed into E.coli DH10Bac,and The recombinant bacmid was contructed.The recombinant bacmid can be used to transfect into sf-9 cells to generate recombinant baculovirus expressing F protein of NA-1.