|Both enzymatic and mechanical lysis can be used for yeast. Enzymatic lysis doesn't require additional lab equipment, but takes longer than mechanical disruption.
Cultures were grown at 30° with constant shaking for 12 hr (in the absence of Cm) or 14 hr (in the presence of Cm). RNA was isolated using the QIAGEN (Ontario, Canada) RNeasy Plant Mini Kit. RNA integrity was measured using an Agilent 2100 Bioanalyzer system and the Agilent RNA 6000 Nano kit (Agilent Technologies). Only those samples that had an RNA integrity number value of >9 were used for cDNA synthesis. First-strand cDNA was synthesized using SuperScript III Reverse Transcriptase (Invitrogen). Reactions were performed using KAPA SYBR Fast qPCR reagents (KAPA Biosystems) and the StepOnePlus qPCR system (Applied Biosystems, Foster City, CA). Data were analyzed using StepOne software (Applied Biosystems). The β-tubulin gene was the internal control for ΔΔCt quantification. The qPCR data for each gene examined were obtained from four biological replicates, each with three technical replicates. Full paper
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