S. Spiegel et al., IMPROVED DETECTION OF PRUNUS NECROTIC RINGSPOT VIRUS BY THE POLYMERASE CHAIN-REACTION, European journal of plant pathology, 102(7), 1996, pp. 681-685
The reverse transcription - polymerase chain reaction (RT-PCR) techniq
ue was used for detection of prunus necrotic ringspot virus in dormant
peach trees which tested negative by ELISA. Total RNA extracted from
bark tissue, using a lithium chloride based method, were used for reve
rse transcription and subsequent amplification of viral sequences. The
PCR product, about 300 base pairs in size, was analyzed by gel electr
ophoresis and visualized by ethidium bromide staining. In some cases,
PCR products were not clearly visible in the stained gel and became di
stinct only following hybridisation with a P-32-labelled virus specifi
c probe. The RT-PCR assay described in this paper is sensitive enough
for detection of PNRSV in dormant woody bark tissue and could be incor
porated into testing protocols during post-entry quarantines for rapid
initial screening of imported budwood and in virus elimination progra
ms.