RNA Preparation from ES Cells

1. Guanidinium lysis buffer: 4 M guanidinium thiocyanate dissolved in 0.1 M Tris-HCl (pH 7.5), filtered through Whatman 3MM paper. Just prior to use, add 2-mercaptoethanol to a final concentration of 1% and sodium lauryl sarcosinate to 0.5% (w/v).

2. 5.7 M cesium chloride (CsCl) dissolved in 10 mM EDTA, pH 7.5, filtered, DEPC-treated, and autoclaved.

3.8 M urea dissolved in 10 mM HEPES, pH 7.0.

4. DEPC H2O prepared in a fume hood by adding 1 mL of diethyl pyrocarbonate (Sigma) to 1 L of dH2O, shake thoroughly and leave overnight before autoclaving.

8. 1:1 Phenol/chloroform.

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