Poster: Biology & Disease Mgmt: Mycology
Development of a multiplex PCR microsatellite marker set for Raffaelea lauricola, and its potential applications
T. DREADEN (1), M. Hughes (2), J. Smith (2) (1) USDA FS, U.S.A.; (2) University of Florida, U.S.A.
Laurel wilt is an emerging disease of trees in the family Lauraceae characterized by systemic wilt and rapid mortality. It is caused by the fungus Raffaelea lauricola and its vector, the redbay ambrosia beetle (Xyleborus glabratus) that was likely introduced to the southeastern United States from Asia. The origin of the US introduction and the population genetics of this devastating pathogen have not yet been characterized. To help facilitate this we developed a panel of microsatellite markers that can be multiplexed, amplify two or more loci in a single reaction, into eight PCRs. The marker set can be used to screen 11 R. lauricola isolates per 96 well PCR plate, eliminating the need to combine samples post amplification. The marker set reduces cost by using conventional hot start Taq, 15ul reaction volumes, multiplexing PCRs and fluorescein-12-dUTP to label PCR amplicons instead of fluorescently labeled primers. Future work will utilize the microsatellite marker set to investigate the genetic structure of R. lauricola within both the US and Asian populations and generate information regarding the number of introductions, possible origins of the introductions into the US and potential for development of new strains.