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C H A P TE R 1
Eukaryotic Target Preparation
Reagent Preparation
5X RNA Fragmentation Buffer
(200 mM Tris-acetate, pH 8.1, 500 mM KOAc, 150 mM MgOAc)
1.
2.
Eukaryotic
The fragmentation buffer is made with RNase-free reagents. Tris-containing solutions
should not be treated with DEPC; however, once H20 has been DEPC-treated and
autoclaved it can be used for making the Tris solution.
Combine the following components to a total volume of 20 mL.
■
4.0 mL 1 M Tris acetate pH 8.1 (Trizma Base, pH adjusted with glacial acetic acid)
■
0.64 g MgOAc
■
0.98 g KOAc
■
DEPC-treated H20 to 20 mL
Mix thoroughly and filter through a 0.2 µm vacuum filter unit. This reagent should be
aliquotted and stored at room temperature.
2.1.7