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Detecting and typing of HFRSV with PCR and biotinylated probes


, : Detecting and typing of HFRSV with PCR and biotinylated probes. Virologica Sinica 9(1): 25-30

By analysis and comparison of nucleotides sequences of HFRSV 76/118 and R-22 strains, three pairs of primers were designed and synthesised. One pair of primer lying in the high homologous region between 76/118 strain and R-22 strain was used as common and outer primers; the other two pairs of primers were in the low homologous region, as the type-specific and inner primers. Using above primers and RT-PCR technique, we measured five strains of HFRSV, 76/118, A9, Chen, R-2, and R-22. When using the outer primers, all of the five strains produced one DNA lane of 300bp;using the field-rat type inner primers all strains but R-22 strain produced one DNA lane of 70 bp and using the home-rat type inner primers, only R-22 strain produced one DNA lane of 70 bp. Part of M fragment cDNA of 76/118 and R-22 strains was used respectively as template, two type-specific biotinylated probed were synthesised by nest PCR technique, the probes were used to hybridization with the RT-PCR products of the five strains, the results showed: RT-PCR technique may be used to the detecting and typing of HFRSV, and had great accuracy, the sensitivity of dot hybridization with biotinylated probe was 1-10 pg of cDNA.

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