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Detection of Andean potato virus X isolates by radioactive and nonradioactive nucleic acid spot hybridization tests

Detection of Andean potato virus X isolates by radioactive and nonradioactive nucleic acid spot hybridization tests

Phytopathology 83(2): 171-176

ISSN/ISBN: 0031-949X

A cDNA probe, pCX61, prepared from the Andean potato virus X (PVX) cp strain was tested against a broad spectrum of Andean PVX isolates grouped in two serotypes: 1) the PVX-A Andean serotype detected only in Peru and Bolivia, which includes the cp and HB strains; and 2) the PVO-O common serotype, which contains isolates serologically similar to those occurring elsewhere in the world. In radioactive nucleic acid spot hybridization tests (R-NASH) of virus in crude sap using the 32P-labeled RNA probe pX61, the PVXA-A isolates showed stronger hybridization signals and a detectability usually from two to six threefold dilution steps higher than those shown for PVX-O isolates. The difference in the detectability of isolates from PVX-A and PVX-O serotypes was similar to that in nonradioactive nucleic acid spot hybridization tests (NR-NASH) with biotinylated DNA probe pX61. However, detectabilty in NR-NAZH was lower than in R-NASH. When biotinylated DNA probes of pX61 (PVX-A-specific) and pPVX19 (PVX-O-specific), prepared from a British PVX isolate of the common European strain-group 3, were compared iN NR-NASH, pPVX19 hybridized much more strongly to most isolates from the PVX-O serotype than to those from the PVX-A serotype. Several PVX-O isolates, mostly from Bolivia, reacted as weakly with pPVX19 as the PVX-A isolates. For R-NASH and NR-NASH tests, virus concentration in crude sap was checked by the double-antibody sandwich form of enzyme-linked immunosorbent assay, confirming that the differences obtained were not because of differences in virus concentration. These results were reconfirmed in R-NASH tests with DNA probes pX61 and pPVX19 using known concentrations of purified RNA from selected isolates. The differences shown between PVX-A and PVX-O isolates stress the importance of using the appropriate probes to detect PVX in breeding programs for resistance and quarantine purposes.

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Accession: 008436898

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