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Detection and quantification of soybean rust (Phakopsora pachyrhizi) spores by quantitative real-time PCR.

Presenter: Todd Steinlage, University of Illinois
Coauthor(s): M.R. Miles2; and G.L. Hartman1,2 1Dept. of Crop Sciences, National Soybean Research Center, University of Illinois, Urbana, IL 61801; 2USDA-ARS and Dept. of Crop Sciences, National Soybean Research Center, University of Illinois, Urbana, IL 61801

An important aspect of an early detection and monitoring system for soybean rust is the reliability of detection and quantification of spores from rain or spore traps. Visual evaluation of rods or slides from spore traps is time consuming and may not be accurate since other look-a-like spores from other species may provide false positives. The most reliable method for identification of soybean rust has been a PCR assay. The main objective of this study was to improve the sensitivity of the QPCR assay for detecting spores from various spore traps. A number of treatments were evaluated including different co-precipitants using fresh and heat-killed spores. The equivalent of DNA from 10 spores was detected at a range of 25 to 27 PCR cycles for fresh spores and a range of 34 to 43 cycles for heat-killed spores, depending upon the test. We have shown that the QPCR method has the sensitivity to detect few spores, but additional research needs to be completed to make this technology more consistent and widely adapted.

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