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Objective To investigate the relationship between the molecular characteristics and phylogenetic evolution of rabies N gene.Methods Saliva samples were collected from rabies cases,and RT-PCR was used to amplify the N gene of rabies virus with the specific primers.The amplifying product of RT-PCR was cloned to pUCm-T vector and transformed into E.coli XL1-Blue and then the blue-white selection,PCR screening and gene sequencing were carried out to identify the positive clones.Finally,ExPASy and other bioinformatics softwares were used to analyze and predict the structure and biological characteristics of the N genome.Results The amplification product of RT-PCR was 1 353 bp,the recombinant plasmid pUCm-T/N was constructed,the whole length of the N gene open reading frame was composed of 1 353 nucleotide residues to code 450 amino acids (20 kinds),the accession number submitted to the Genbank was HM756692,its sequence homology of nucleotides and amino acids compared with the vaccine strain CTN-1-V was 90% and 99% respectively.The evolutionary analysis showed that the isolated strain belonged to genotype I with certain geographic regionality.Conclusion The characteristics investigation and bioinformatics analysis of Hunan0806 N gene will provide fundament data to reveal the significance of the N gene characteristics for rabies epidemiology and its prevention & control.
Objective To investigate the relationship between the molecular characteristics and phylogenetic evolution of rabies N gene. Methods Saliva samples were collected from rabies cases, and RT-PCR was used to amplify the N gene of rabies virus with the specific primers. Amplifying product of RT -PCR was cloned to pUCm-T vector and transformed into E. coli XL1-Blue and then the blue-white selection, PCR screening and gene sequencing were carried out to identify the positive clones. Finaally, ExPASy and other bioinformatics software were used to analyze and predict the structure and biological characteristics of the N genome. Results The Amplification product of RT-PCR was 1 353 bp, the recombinant plasmid pUCm-T / N was constructed, the whole length of the N gene open reading frame was composed of 1 353 nucleotide residues to code 450 amino acids (20 kinds), the accession number submitted to the Genbank was HM756692, its sequence homology of nucleotides and amino acids compared with the vaccine strain CTN-1-V was 90% and 99% respectively. The evolutionary analysis showed that the isolated strain belonged to genotype I with certain geographic regionality. Confluence The characteristics investigation and bioinformatics analysis of Hunan0806 N gene will provide fundament data to reveal the significance of the N gene characteristics for rabies epidemiology and its prevention & control.