Figure 1.

Efficient extraction of yeast RNA by heating yeast cells in the presence of formamide and EDTA. (A) Effects of temperature and additives on the efficiency of RNA extraction by formamide. A total of 2.5 OD₆₀₀ units of BY4741 cells (culture at OD₆₀₀ = 5.0) was resuspended in formamide (40 μl per 1.0 OD₆₀₀ unit of cells), incubated for 5 min at the indicated temperature, vortexed for 10 s and centrifuged for 1 min at 12,500 rpm in a microcentrifuge. Equal volumes of each supernatant were separated on a 1% agarose gel and RNA was stained with SYBR Gold. Incubations at 65°C were either in pure formamide or in formamide with the addition of 10 mM EDTA, 20 mM TCEP, 1 × HT electrophoresis buffer, or 0.2 × SSC. (B) FAE extraction preserves RNA integrity. Electrophoretic analysis of 2.5 μg RNA prepared from the same culture by one-step FAE extraction (lane 1), FAE extraction with additional vortexing of cells for 8 min using 0.5 mm glass beads (lane 2) and RNAsnap™ (lane 3). RNA was transferred to a membrane and analyzed by northern hybridization with a probe that detects 25S rRNA.