Profiling and quantifying differential gene transcription in Phytophthora infestans prior to and during the early stages of potato infection.

Phytophthora infestans, the causal agent of potato and tomato late blight, produces several different cell types prior to and during the early stages of potato infection. All of these cell types can be easily generated and studied in the absence of the host plant and so form the basis for developmental stage-specific gene discovery. We have used amplified fragment length polymorphism (AFLP)-based mRNA fingerprinting (cDNA-AFLP) to identify 64 transcripts that appeared to be up-regulated in germinating cysts but not in vegetative mycelium. These transcripts included representatives of most major classes of heat shock proteins: hsp60, hsp70, hsp90, and hsp100. Real-time RT-PCR was used to quantify the expression of 18 transcripts originating from germinating cysts, relative to the constitutively expressed actB gene, in vegetative mycelium, germinating cysts, and at three time-points post-inoculation of potato cultivar Bintje (15, 48, and 72h). All of the transcripts were up-regulated in germinating cysts, and 12, including hsp70, hsp80-2, and hsp90, were found also to be up-regulated in planta. This is the first report of the application of real-time RT-PCR to the relative quantification of plant pathogen gene expression during the early stages of infection.