TmHox gene expression analysis in Talaromyces marneffei

Gene expression analysis from human fungal pathogen Talaromyces marneffei strain ATCC200051. To prepare <i>T. marneffei</i> in different growth forms, the conidia were harvested and 1x10⁸ conidia/ml were inoculated in a 50-ml Sabouraud’s dextrose broth (SDB). Cultures were incubated at 25°C (mycelium phase) and 37°C (yeast phase) with continuous shaking at 200 rpm. After 72 hours, the stable mycelium and yeast cultures were collected by centrifugation at 4°C, 7,000 rpm for 30 min. For the phase transition experiment, 1x10⁸ conidia/ml were inoculated in a 5-ml SDB and incubated at 25°C (mycelium phase) and 37°C (yeast phase) with continuous shaking at 200 rpm. Cultures were harvested at 30-min, 1-hr, and 2-hr by centrifugation at 4°C, 7,000 rpm for 30 min. RNA were extracted by using TRIzol® reagent, treated with DNase I, and converted to cDNA . Quantitative real-time PCR was performed using the SYBR Green qPCR mix (Thunderbird SYBR Green Chemistry, TOYOBO. Osaka, Japan). An actin gene was included as a reference gene (internal control).