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Star Republic: Guide for Biologists

Formaldehyde-Agarose Gels

Edited by Chang Zhu
1. Prepare 10X MOPS buffer (200 mM 3-(N-morpholino)propanesulphonic acid (MOPS), 80 mM sodium acetate, 10 mM EDTA, pH 7.0). Make from DEPC-treated stocks of 1 M MOPS (pH 7.0), 0.5 M EDTA, 3 M sodium acetate, and ddH2O.

2. Clean gel box, tray, and comb throughly with detergent (or soak in 0.2 M NaOH for 10 minutes) and then rinse with distilled water.

3. To prepare 0.7%-1.5% agarose gel, melt 2% agarose in RNase-free ddH2O. Cool down to 60 C.

4. For 100 ml formaldehyde-agarose gel, add 20 ml 37% formaldehyde, 10 ml 10X MOPS, appropriate amount of 2% agarose, nuclease-free ddH2O, and 5 ml 10 mg/ml ethidium bromide (EB). EB may be omitted in the gel but added in the loading buffer.

5. Cast the gel.

6. Heat the sample in 1X RNA loading buffer to 65-70 C for 5 minutes.

6. Run the gel in the 1X MOPS buffer.