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Development of routine RT-PCR tests for certification of fruit tree multiplication material


  • Kummert, J. , Vendrame, M. , Steyer, S. & Lepoivre, P. (2000). Development of routine RT-PCR tests for certification of fruit tree multiplication material. Bulletin OEPP, 30: (3/4),
Type Journal Article
Year 2000
Title Development of routine RT-PCR tests for certification of fruit tree multiplication material
Journal Bulletin OEPP
Recnumber 8
Volume 30
Issue 3/4
Endnote Keywords certification|plant pathogens|planting stock|polymerase chain reaction|pome fruits|stone fruits|techniques|
Abstract Current developments in certification procedures for propagating material require the availability of rapid, sensitive, reliable and user-friendly detection protocols applicable for routine testing. Our research concerns the possible use of reverse transcriptase-polymerase chain reaction (RT-PCR) for the detection of the pathogens listed for virus-tested pome and stone-fruit propagating material in Belgium. Although RT-PCR satisfies the need for rapidity and sensitivity, the usual protocols relying on the use of purified nucleic acid preparations as template and ethidium bromide-stained agarose gels for detection are not appropriate for routine use. We therefore first optimized the parameters and cycling conditions of the RT-PCR reactions to allow direct use of crude extracts of either leaf or bark material as a template. Sandwich hybridization between a covalently linked capture probe and a biotinylated detection probe was then used for the detection of the specific amplicons. These assays have the sensitivity and specificity of the RT-PCR, enhanced by sandwich hybridization with specific probes, and ease of sample preparation and detection of the amplicons. They make it possible to analyse a great number of samples and are thus well adapted for routine quality-control testing of propagating material.
Notes Cited Reference Count: 10 ref. Journal article, Conference paper English
Author address Faculte Universitaire des Sciences agronomiques, Unite de Phytopathologie, 2 Passage des Deportes, B-5030 Gembloux, Belgium.
Fichier
Lien ://20013080614
Authors Kummert, J. , Vendrame, M. , Steyer, S. & Lepoivre, P.