This project will investigate the feasibility of producing a diagnostic chip for the simultaneous detection of all the plant pests/pathogens present in the European Union Plant Health Directive, using quarantine potato pests as a model system. The project will establish methods for detection of quarantine pathogens in plant tissue using gene chip technology. Up to 30 000 DNA probes (sequences from each of the organisms to be tested in one assay) can be arrayed onto a microscope slide - the 'gene chip'. The chip can be exposed to fluorescently labelled nucleic acid from the sample and fed into a micro-array reader to reveal if any of the pathogens were present. The project covers the whole process from preparing target genes, generating gene chips, preparing samples, hybridisation of samples, analysis of results and finally through to quarantine diagnosticians evaluating the technology.
The objective of the project is to establish methodologies for the direct detection of quarantine pathogens of potato in suspect plant samples using gene chip technology. The gene chip consists of a glass microscope slide with the gene sequences from each of the organisms that need to be detected in a single assay arrayed on its surface. Methods will be established for sample extraction, sample labelling, hybridisation and data capture. The two most important characteristics of a diagnostic protocol will be compared with traditional methods, i.e. sensitivity and specificity. The end point of the project is a 'ring test' where, following a training course to facilitate technology transfer, the technology developed will be evaluated by quarantine diagnosticians.
1) To identify existing, or generate new, specific nucleic acid probes for each of the quarantine potato organisms, except Colorado beetle and three IAI fungi.
2) To array the probes from objective (1) onto slides at a density of 900 spots per slide or greater, and stabilise chip suitable for mailing to collaborators.
3) To develop a micro-array extraction, labelling and hybridisation protocol for the simultaneous detection of DNA and RNA organisms in infected/infested potato leaf/tuber tissue.
4) To determine limits of senstivity and scificity of the prototype gene chip.
5) To array out final optimal probes, to generate DIAG CHIPS for validation.
6) To validate the protocol by submitting to a ring test by quarantine testing stations from at least four EU states.
ARABLE CROPS, CROP PESTS AND DISEASES, GENOMICS, QUANTITATIVE APPROACHES AND MODELLING
Scientist responsible for the project
Mr STEPHEN HILL
YO41 1LZ York
United Kingdom (The) - GB
Phone: +44 1904 462227
Fax: +44 1904 462250
||CENTRAL SCIENCE LABORATORY, MINISTRY OF AGRICULTURE, FISHERIES AND FOOD
||01 December 2001
||1 452 408 €
|Total EC contribution
||980 783 €
|Web address of the project