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Star Republic: Guide for Biologists

PCR cleanup --- 96-well

1. Add 180 ml of 150 mM potassium acetate (pH 4.8) and 5.3 M guanidine-HCl to each well of the PCR reactions.

2. Place Millipore glass filter plate on the vacuum manifold or a collection plate.

3. Wet the filters with 20 ml of 150 mM potassium acetate (pH 4.8) and 5.3 M guanidine-HCl.

4. Transfer the PCR mix to the filter plate.

5. Apply vacuum or centrifuge at 3,000 rpm. DNA will bind to the glass filters.

6. Wash the filters with 200 ml of 80% ethanol. Apply vacuum or centrifuge at 3,000 rpm.

7. Repeat 80% ethanol wash 3 times.

8. After the final wash, place the filter plate on a collection plate (discard flow through in the collection plate first) and centrifuge at 3000 rpm for 5 minutes to remove residual ethanol.

9. Place filter plate on top of a clean 96-well microtiter plate. Align wells and tape the two plates together. Add ~50 ml of ddH2 or TE to each well. Let stand for 1 minute at room temperature. Centrifuge at 3,000 rpm for 5 minutes to elute DNA.