For induction, ethanol-dissolved BOA (50 µg ml-1 final conc.; 1% ethanol) was added to cultures, while only ethanol (1%) was added for the BOA-unexposed treatment. After 2 hr incubation, the fungal growth was harvested by vacuum filtration, flash frozen in liquid nitrogen, and stored at -80C until ready for extraction.
Growth protocol
Wild-type F. verticillioides FRC M-3125 was grown for three days in PDB (50 ml) at 200 rpm in the dark at 27C. The fungus then was transferred (1 ml) to replicate flasks of fresh PDB (50 ml) and grown for three more days to synchronize cultures.
Extracted molecule
total RNA
Extraction protocol
Samples were ground with liquid nitrogen, and RNA was extracted with Ambion PureLink RNA Mini Kit (Life Technologies, Carlsbad, CA, USA) using homogenizers, followed by treatment with Turbo DNase (Life Technologies), all according to manufacturer’s instructions. RNA integrity was evaluated on an Agilent 2100 Bioanalyzer. Double stranded cDNA was generated using Roche cDNA Synthesis System (Indianapolis, IN, USA), again according to instructions.
Label
Cy3
Label protocol
Labeling was performed by NimbleGen Systems Inc., Madison, WI USA, following their standard operating protocol.
Hybridization protocol
Hybridization was performed by NimbleGen Systems Inc., Madison, WI, USA following their standard operating protocol.
Scan protocol
Scanning was performed by NimbleGen Systems Inc., Madison, WI USA, following their standard operating protocol.
Data processing
The normalized .calls files provided by NimbleGen (representing normalized gene expression values and accession numbers for each gene interrogated by the array) were imported into ArrayStar v.11 for comparative analysis.
