Fluorescent in situ RT-PCR to visualise the expression of a phosphate transporter gene from an ectomycorrhizal fungus.

ABSTRACT

Expression of a mycorrhizal fungal-specific phosphate (P) transporter gene (HcPT1) was studied in mycelium of the ectomycorrhizal fungus Hebeloma cylindrosporum, by in situ reverse transcriptase polymerase chain reaction using amplification of complementary DNA sequences. The expression of HcPT1 was visualised under two different P treatments. Mycelium was transferred to liquid medium with or without P and incubated for 5 days. Under P starvation, mycelium growth and vitality was reduced and the expression of HcPT1 up regulated. Enzyme-labelled fluorescent substrate was used to detect gene expression in situ with epi-fluorescence microscopy and to visualise it at the level of the individual hyphae both in starved and non-starved hyphae. Up-regulation of HcPT1 was observed as a more intense fluorescent signal and from the larger proportion of hyphae that showed expression.