To identify the genome-wide transcriptional changes that occur throughout sexual developmen of C. neoformans, we conducted a time-course microarray experiment spanning six developmental stages to generate a temporal expression pattern for each known gene. C. neoformans a x α co-cultures were grown under sexual development conditions (V8 agar), and RNA was harvested at 0.5, 6, 12, 24, 48, and 72 hours post-mixing. Time points were chosen based on microscopic identification of cell types of interest: fusants, filaments (early and late), basidia, and spores (Figure 1B). Microarray hybridizations were conducted in a loop design, with each sample serving as a reference for the following time point in the experiment (e.g. 0.5 hours vs. 6 hours, 6 hours vs. 12 hours, etc).
Overall design: The experiment spanned six time points of development: 0.5 hours, 6 hours, 12 hours, 24 hours, 48 hours and 72 hours. Hybridizations were conducted in a loop design, with each sample serving as the reference for the consecutive time point - resulting in 5 experimental comparisons: .5hr vs 6 hr, 6 hrs vs 12 hrs, 12 hrs vs 24 hrs, 24 hrs vs 48 hrs, and 48 hrs vs 72 hrs. Each comparison consistent of duplicate hybridizations with dye swaps. Arrays contain duplicate genomes, resulting in eight-fold coverage within each comparison. (total of 20 hybridizations)
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