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The Full Length Genome Sequence of a Newcastle Disease Virus (ZJ1) of Goose Origin and the Construction of a Transcription Vector Containing the cDNA of Its Entire Genome

Author: HuangYong
Tutor: LiuXiuZuo
School: Yangzhou University
Course: Preventive Veterinary Medicine
Keywords: goose Newcastle disease virus genome Nucleocapsid protein transcription vector
CLC: S852.65
Type: PhD thesis
Year: 2003
Downloads: 204
Quote: 13
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Abstract


Newcastle disease virus(NDV) is one of the most important animal pathogens of poultry, because it can infect many avian species and often causes severe losses in poultry industry. Up till now, the complete genomic sequences of NDV strains of B1, Lasota, Clone-30, Beaudette C and HB V4 have been published and/or released on GenBank, they all have the same length of 15186 nucleotides. The completion of the genomic sequence of NDV leads to the development of a reverse genetic system for this virus, opening a new approach in gene research and vaccine development.Since 1997, a novel disease entity in the goose with clinical signs and lesions resembling those of ND in chickens has been reported from regions in South and East China. Virulent ND viruses could be invariably isolated from diseased geese in field outbreaks. Several NDV isolates of goose origin have been characterized pathotypically and genotypically in recent years, hi this study, we determined the complete genome nucleotide sequence of an NDV of goose origin(ZJl) which consists of 15192 nt, 6 nt longer than those of other five NDV strains, and this additional six nt motif is located in the non-coding region of NP gene. Other 17 NDV strains of different genetic and epidemiological background were also searched for the existence of this motif, hi addition, a transcription vector containing the full length cDNA from ZJ1 genome and three expression plasmids for NP, P and L protein were also constructed.1 The entire genomic sequence of NDV strain ZJ1 and its comparison with those of other NDVsBy comparing the gene sequences of different NDVs released on GenBank, nine pairs of primers were designed on the basis of the sequences of conserved regions with high homology among NDVs. Each fragment amplified by RT-PCR has overlapping sequences with neighbouring fragments hi NDV genome. A total of rune overlapping viral cDNA fragments spanning the whole internal region of ZJ1 genome were obtained and sequenced. The sequences of the 3’ - and 5’ - terminal ends of viral RNA genome were determined by rapid amplification of cDNA ends (RACE). The entire genome sequence of ZJ1 compiled from the nine overlapping viral cDNA fragments and sequences of both 3’ and 5’ ends is 15192 nt, six nt longer than those of other 5 NDV strains (Lasota, Bl, Clone-30, Beaudette C and HB V4). This six nt motif is located in the untranslated region of nucleoprotein (NP) gene between the corresponding positions 1646nt and 1647nt of the genome of Lasota Strain.Similar to the genomic structure of the other five NDVs, ZJ1 genome comprises six genes, from the 3’-terminus to the 5’-terminus, which encode the nucleocapsid protein (NP), phosphoprotein (P), matrix protein (M), fusion protein (F), haemaggluti and successively. The lengths of NP, P, M, F and L of ZJ1 in amino acid residues are the same as those of the other five NDVs, whereas the HN of ZJ1 is 571 amino acid long sharing the typical characteristic of virulent NDVs. In ZJ1 genome, the open reading frame(ORF) coding for the small hydrophobic protein (SH) is absent, whereas the RNA editing site (AAAAA- GG I G) is located at position 484 of the P geneWith the exception of the additional six nt motif in the corresponding position 1646nt~1647nt of lasota strain, the positions of start and end, and the length of each gene of ZJ1 share high degree of identity with those of the other five NDVs. The lengths of the intergenic regions NP-P, P-M, M-F, F-HN and HN-L in ZJ1 genome are lnt(T), lnt(T), lnt(T), 31nt and 47nt respectively. The sequences of NP-P, P-M and M-F junctions of ZJ1 and the other five NDVs share high degree of identity, while the sequences of F-HN and HN-L junctions of ZJ1 have low degree of identity with those of the other five NDVs.Comparing the coding sequences of six genes of ZJ1 and the other five NDVs, thenucleotide sequence identities among other five NDVs are high, whereas those between ZJ1 and the other five NDVs are relatively low.The leader and trailer sequences of ZJ1 genome are 55nt and 114nt lon

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CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > Animal Medicine ( Veterinary Medicine) > Basic Veterinary Science > Animal Microbiology ( Veterinary Microbiology, ) > Livestock Virology
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