Cloning symbiosis-related cDNAs from eucalypt ectomycorrhiza by PCR-assisted differential screening

As part of a project to identify symbiosis-related genes, we report here a simple differential screening procedure for isolating up- and down-regulated fungal transcripts from a cDNA library of the developing Eucalyptus globulus-Pisolithus tinctorius mycorrhiza. cDNA inserts of randomly selected λZAP plaques were amplified by PCR and separated by agarose gel electrophoresis. The PCR-amplified cDNA samples were then screened by Southern blotting, using radiolabelled-cDNA probes of high specific activity. We have applied this method to fungal transcripts that are differentially expressed in ectomycorrhizas during the early stages of development. We estimate that about 50 % of the fungal mRNA population is regulated by development of the symbiosis; several up- and down-regulated cDNAs have been isolated for further analysis.