Application of semi-nested PCR for the genotype analysis of rubella virus

Abstract

Objective To identify the genotypes of rubella virus (RV) that had not been isolaed by cell culture, and to understand comprehensively the molecular epidemiological features of RV in Hebei province. Methods Throat swab samples which were positive by real time fluorescence quantitative PCR were cultured with three times of passaging and were tested for nucleic acid of RV. The samples with negative results after culture were selected for amplification by the semi-nested PCR and the target fragments were sequenced. Results Among the selected eleven RV throat swab samples, six were genotype 2B and five were genotype 1E. All these genotypes were highly homologous to their correspondent reference strains. No mutations were found in important sites, for example glycosylation sites, epitopes, hemagglutination inhibition and neutralization sites. Conclusions Throat swab samples can be genotyped by semi-nested PCR. This will enrich the gene sequence bank of RV and provide basis for comprehensive understanding the molecular epidemiology of RV in Hebei province. Key words: Rubella virus; Genotype; Semi-nested PCR; Molecular epidemiology.