Day 5 began with a lecture by OSU Ph.D. student Nagendra Subedi on the theory and practice of Real-Time PCR. Real-time PCR technology has moved from research labs into diagnostic applications for numerous plant pathogens. The technique requires less time and labor to perform than traditional PCR, and allows quantitation of the diagnostic target in a sample. DNA was extracted on Day 4 from tomato plants suspected to be infected with Clavibacter michiganensis subsp. michiganensis (Cmm), the causal agent of bacterial canker.
Bacterial canker of tomato – note typical marginal necrosis.
“Bird’s-eye” lesions typical of bacterial canker on tomato fruit.
DNA extracts were tested for Cmm by both conventional (endpoint) and Real-Time PCR (TaqMan). Results were positive for all suspected samples by RT- PCR, a few were missed in endpoint PCR. Participants also tested the putative Cmm samples with DNAble tests kits – DNAble kits are based on isothermal DNA amplification technology.
Dr. Sally Miller presented a case study for detection and diagnosis of Ralstonia solanacearum, a potentially devastating disease of many crops in the subtropics and tropics. All of the diagnostic techniques in practice in the short course so far are used in identification of this pathogen.