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Transcriptome evolution in the yeast cell cycle

How to make DEPC-treated water and Tris Buffer

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created Sat, 09 Sep 2006 17:39:00 | modified Sat, 09 Sep 2006 17:39:00
Abstract: Solutions (water and other solutions) should be treated with 0.1% DEPC. DEPC is a strong, but not absolute, inhibitor of RNases. It is commonly used at a concentration of 0.1% to inactivate RNases on glass or plasticware or to create RNase-free solutions and water. DEPC inactivates RNases by covalent modification.
Source: Qiagen
  1. Add 0.1 ml DEPC to 100 ml of the solution to be treated and shake vigorously to bring the DEPC into solution.
  2. Let the solution incubate for 12 hours at 37°C.
  3. Autoclave for 15 minutes to remove any trace of DEPC.

Notes: 1. How to make DEPC-treated Tris buffers: DEPC will react with primary amines and cannot be used directly to treat Tris buffers. DEPC is highly unstable in the presence of Tris buffers and decomposes rapidly into ethanol and CO2. When preparing Tris buffers, treat water with DEPC first, and then dissolve Tris to make the appropriate buffer. Trace amounts of DEPC will modify purine residues in RNA by carboxymethylation. Carboxymethylated RNA is translated with very low efficiency in cell-free systems. However, its ability to form DNA:RNA or RNA:RNA hybrids is not seriously affected unless a large fraction of the purine residues have been modified. Residual DEPC must always be eliminated from solutions or vessels by autoclaving or heating to 100°C for 15 minutes.

Protocol Online http://www.protocol-online.org


DEPC - Explosive

Diethyl pyrocarbonate (DEPC) is commonly used in molecular biology. DEPC is moisture sensitive and hydrolyzes to CO2 and ethanol upon exposure to moist air or water. Bottles of DEPC will absorb moisture upon being opened; release of CO2 during subsequent storage may lead to pressure buildup and spontaneous explosion. Such an incident recently occurred in a lab at another university. Spontaneous explosion of new unopened bottles can also occur.

Every bottle of DEPC, whether new or previously opened, should be considered an explosive hazard. DEPC should be purchased in small quantities and used as soon as possible after purchase. DEPC should be stored tightly capped and refrigerated, with desiccant, in the original metal shipping containers. These may be placed inside larger metal receptacles for added protection. Allow DEPC to equilibrate to room temperature before handling. Wear goggles and protective clothing, and open bottles behind a shield in a chemical fume hood.

Adapted from “Lab Safety Spectrum,” McGill University