Magnetic Capture Hybridization and Real-Time Multiplex PCR for the Detection of Seedborne Pathogens

NON-TECHNICAL SUMMARY: Seeds are internationally traded commodities, that represent a potential weak link in US plant biosecurity. Seeds may become infested with plant pathogens, and serve as vectors to introduce organisms to the US. Current seed detection lacks the sensitivity to completely exclude seedborne pathogens. The PCR is a powerful detection tool that has the potential to improve seed health testing. The goal of this project is to develop magnetic capture hybridization (MCH) and real-time PCR to improve the sensitivity, and efficiency of pathogen detection in seed. By training private and public sector seed pathologists to develop and implement PCR- based assays, we will increase the chances of detecting infested seed lots and thereby improve plant biosecurity. <P>

APPROACH: The objective of the proposed research is to develop a sensitive, specific and efficient assay for simultaneously detecting multiple pathogens in seeds. This will require the development of specific PCR primers, hybridization capture and TaqMan probes. Once the multiplex MCH real-time PCR assay has been developed it will optimized and comparatively evaluated. By designing similar assays for different host-pathogen combinations we hope to demonstrate the universal applicability of this system. We also propose to develop an outreach program whereby diagnosticians and seed pathologists can be trained to develop and implement MCH real-time PCR assays to improve their ability to accurately detect key pathogens in seed.