Simultaneous RT/PCR detection and differentiation of arabis mosaic and grapevine fanleaf nepoviruses in grapevines with a single pair of primers

Abstract

The movement protein genes from several isolates of ArMV and GFLV of different geographical origins were amplified by RT/PCR using degenerate primers, cloned and sequenced. A single pair of degenerate primers was designed from these sequences to allow the simultaneous amplification of parts of the movement protein genes of ArMV and GFLV. Their use in an immunocapture-RT/PCR for the detection of ArMV or GFLV in infected grapevines proved to be ten times more sensitive than the corresponding ArMV or GFLV ELISA tests. A Sph1 restriction site found in the sequences corresponding to the amplified products from the GFLV isolates, but not in the amplified products from the ArMV isolates, allowed the differentiation between ArMV and GFLV in the infected grapevines by a Sph1 restriction digestion of the amplified products.