|SAITO, SEIYA - Cornell University|
|WILCOX, WAYNE - Cornell University|
Submitted to: American Phytopathological Society Abstracts
Publication Type: Abstract Only
Publication Acceptance Date: 3/15/2012
Publication Date: 5/1/2012
Citation: Saito, S., Cadle Davidson, L.E., Wilcox, W. 2012. Comparative analysis of techniques for detection of quiescent Botrytis cinerea in grapes by quantitative PCR. American Phytopathological Society Abstracts. Phytopathology 102:S.
Technical Abstract: Quantitative PCR (qPCR) can be used to detect and monitor pathogen colonization, but early attempts to apply the technology to quiescent Botrytis cinerea infections of grape berries identified some specific limitations. In this study, four DNA extraction methods, two tissue-grinding methods, two grape (Vitis vinifera) organs, three probe sets and two enzymes were compared in order to improve the sensitivity of B. cinerea detection in grapevine. Furthermore, duplex qPCR for concurrent detection of B. cinerea and V. vinifera DNA was developed as a check against false negative assays, and to establish a Pathogen Coefficient (PC) that addresses variability among samples caused by DNA quality, PCR efficiency and pipetting error. The resulting optimized qPCR technique was highly sensitive, providing a Ct value less than 33 for as little as 0.001 ng of B. cinerea DNA, and could be applied as a tool to monitor quiescent B. cinerea infections in vineyards.