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Star Republic: Guide for Biologists

Plasmid prep --- 96-well

Edited by Chang Zhu
1. Grow cells in 96 deep-well block.

2. Pellet cells by centrifugation at 2,000 rpm for 5 minutes.

3. Pour off the supernatant. Drain off the rest of supernatant by inverting the block on paper towels or by aspiration.

4. Cover the plate and freeze at -80 C for 15 minutes.

5. Thaw the plate at room temperature.

6. Resuspend the cell pellet in 100 ml P1 + 0.1 mg/ml RNase A. Vortex at maximum speed for 2 minutes.

7. Add 100 ml P2, vortex at low speed for 10 seconds. Let stand on ice for 5 minutes. Alternatively, add 100 ml P2, shake by hands to mix, and then leave at room temperature for 1 hour.

8. Add 100 ml P3, vortex to mix. On ice for 5 minutes.

9. Centrifuge at 3,000 rpm for 30 minutes.

10. Remove 200 ml supernatant and add to a second deep well plate containing 200 ml isopropanol in each well.

Precipitate in the deep well plate by centrifuging at 3,000 rpm for 30 minutes. Alternatively, transfer 200 ml to a microtiter plate, precipitate DNA. Discard the supernatant and transfer the remaining 200 ml to the microtiter plate with pellets, precipitate DNA.

11. Wash with 70% ethanol. Decant the supernatant, drain inverted on a paper towel. Dry under vacuum or air dry.

12. Resuspend in 100 ml ddH2O or TE.

See also Alkaline lysis plasmid miniprep.