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Construction of genomic full-length cDNA of SARS coronavirus BJ01 strain and identification of its biological characteristics



Construction of genomic full-length cDNA of SARS coronavirus BJ01 strain and identification of its biological characteristics



Wei Sheng Wu Xue Bao 46(6): 922-927



Severe acute respiratory syndrome (SARS) is an important emerging infectious disease which caused by SARS coronavirus (SARS-CoV), and the study of its pathogenesis is needed for the treatment and prevention of this disease. To study the pathogenesis of SARS-CoV using reverse genetics technology, by in vitro ligation using 7 contiguous cDNAs that span the entire genome of the SARS-CoV BJ01 strain, a genomic full-length cDNA was assembled, then using T7 RiboMAX Large Scale RNA Production Systems with the genomic full-length cDNA as template, the RNA transcript was attained. The typical SARS-CoV-resulted cell pathogenic effects were observed when RNA transcript was electroporated into Vero E6 cells. The results of RT-PCR and sequencing of the rescued virus showed that it originated from transcript which derived from the full-length cDNA construct. Rescued virus-infected cells were detected by indirect fluorescent antibody staining demonstrated that it can specifically reaction with SARS-CoV. By CPE method and plaque assay, the titers of the rescued virus and wild-type virus were assessed, which demonstrated there are no significant difference between the viruses and they have similar biological characteristics. Construction of the genomic full-length cDNA of SARS-CoV BJ01 stain successfully and study of the biological characteristics of the rescued virus will provide a useful tool serving for the discovery of molecular pathogenesis and development of candidate vaccines against SARS-CoV.

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Accession: 048642233

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PMID: 17302155


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