| To gain insight to the interaction between NDV and host cells, the relationships between structure and function,the difference between the velogenic strains (such as F48E9) and lentogenic strain, and to develop a new live attenuated virus vaccines or a new viral vector with NDV,we have cloned the full length cDNA of NDV F48E9 genome.Nine pair primers were designed according to the sequence of NDV La Sota strain and partial F48E9 strain.Eight cDNA clones spanning the genomic sequence of NDV strain F48E9 were generated from viral RNA by RT-PCR using above NDV specific primers. The PCR fragments were cloned into pMD18-T vector,after restriction endonuclease analysis ,the genomic sequence of NDV strain F48E9 cDNA was determined.Then another five pairs and half primers were designed according the sequence of the above PCR products. The fragments were subcloned into a low copy transcription vector (pX8dT) between the T7 RNA polymerase promoter and autocatalytic hepatitis delta virus ribozyme.The result showed that genome of NDV F48E9 strain comprises 15192 nt,which was equal to ZJl strain and 6 nucleosides longer than that of La Sota,V4,Bl and Clone30. There were six major open reading frames in genome of F48E9 strain which encode the viral NP,P,M,F,HN and L protein,respectively. It was interested that there was an extra six nucleosides insertion between 1647-1652nt (according to the genomic sequence of La Sota strain), and the sequence were CCCCCC in F48E9 strain ,and TCCCAC in ZJ1 strain .In order to test if insertion of this six nucleosides is related with the virulence of ND, two primers were designed to amplify the same fragment of another ten NDV strains. The result of sequence comparison of 16 strains showed that the six nucleoside was absent in lentogenic strain.This suggested that the six nucleosides insertion might have relationship with the NDV virulence. Compared with all known sequence of NDV.there was a special sequence (5'TCTCTCTCCTCTCTCCTCC3') in the genomic cDNA of NDV F48E9 strain. We also did not find the same sequence in the Genbank. So we think that it is a F48E9 strain specific, and the function of this sequence remain to investigate further more.The nucleoside identity of the genomic sequence of strain F48E9 to that of La Sota,V4,B1,ZJ1 and Clone 30 strain was found to be 87.3%,88.8%,87.3%,85.4% and 87.7%. On the amino acid sequence level, an identity was 90.8-99.6%(NP), 79.5-99.5%(P). 89-99.7%(M), 87.7-99.8%(F), 88.6-99.5%(HN) and 92.2-99.6%(L) for this six strain. However, we found a reigon of 30 amino acids in L protein (aa 1287-1318 of the respective L protein) of F48E9 which is different from the La Sota,ZJ1,B1 and Beaudette-C and having a higher homology with V4 and Clone30 strain.All the result showed that NDV F48E9 strain has its own speciality compared with other five NDV strains,and there were many difference between velogenic strains and lentogenic strains. So the infectious cDNA of rnesogenic strains and lentogenic strain was far from enough to understand the replication, pathogenicity of NDV and the interaction between NDV and host cells,and the infectious cDNA of velogenic strains ( eg.. F48E9 ) was required to explain the relationships between structure and function.Candidate : YanLihui Speciality: Prevention Veterinary Science Supervisor: Prof. Liu Baoquan Prof. Cao Dianjun...
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