Bio-Rad Aurum™ Total RNA Fatty and Fibrous Tissue Kit - Manual (Page 12)

8
(DEPC) to inactivate RNases. DEPC is an efficient, strong, and nonspecific
RNase inhibitor that is usually used at a concentration of 0.1%
• To prepare a 0.1% (v/v) solution of DEPC-treated water, add 1.0 ml of
liquid DEPC per 1 L of water. Incubate the solution at 37°C for 1 hr while
mixing thoroughly. Autoclave the treated water to remove the DEPC
• Warning: DEPC is suspected to be a carcinogen and should be handled
with care. Always use gloves and open under a fume hood
Preparation of 10 mM Tris, pH 7.5, for DNase I Reconstitution
• To prepare 50 ml of 10 mM Tris, pH 7.5, add 60.6 mg of Tris (catalog #
161-0716) to 45 ml of DEPC-treated water. Mix until Tris is completely
dissolved. Adjust the pH of the solution by dropwise addition of 6N HCl.
Once the pH is adjusted to 7.5, add more DEPC-treated water to make a
final volume of 50 ml
Note: DEPC is destroyed by primary amines (e.g., Tris). If a solution
containing a primary amine will be DEPC-treated, omit the amine in
preparing the solution. Perform the DEPC treatment as described above
and add the amine to the autoclaved solution once the solution has
cooled
Preparation of 70% Ethanol
• To prepare 100 ml of 70% ethanol, add 70 ml of 95–100% ethanol to 30
ml of DEPC-treated water. Mix well before use.
Maintaining an RNase-Free Environment
• Although the components of this kit are provided free of contaminating
ribonucleases, great care must be taken not to contaminate the solutions
or the RNA binding columns. Gloves should always be worn when
handling RNA and should be changed frequently. Proceed through the
RNA isolation as quickly as possible with care
• Solutions that are prepared by the user should be treated with DEPC to
inactivate RNases as described above.
10001298B:4110133A.qxd 1/16/2009 2:40 PM Page 8