Polymerase chain reaction and non radio labelled DNA probe for detection of bluetongue viruses

Abstract

Bluetongue is economically important viral disease of ruminants, Keeping this in mind, nucleic acid based assays, viz, RNA-PAGE, RT-PCR and non radio labelled DNA probes,were evaluated for detection of BLU virus in infected BHK 21 cell culture, The RNA-PAGE revealed differences in migration pattern of RNA segments between serotypes I and 18 while there were additional segments in H strain of BLU virus serotype L To develop group specific RT-PCR, segment
6 (NS I gene) was targeted. The primers used, successfully amplified the desired partial length product of 274 bp in 30
PCR cycles. Since the primers produced the expected 274 bp products using 2 different strains of BLU virus serotype I
and one isolate of serotype 18, it was assumed that the amplification was group specific. The PCR product (274 bp)
obtained was labelled with digoxigenin (DIG) and evaluated for detection of the BLU viruses grown in cell culture. The
DIG labelled 274 bp DNA probe reproducibly detected cell culture grown BLU viruses irrespective of the strain/serotype.
This study clearly demonstrated the potential application of nucleic acid based assays for group specific detection of BLU
viruses.