Preparation of DNA Using Concert High Purity Plasmid Mini Prep System

1. Cell Harvesting: Pellet 1 to 5 ml of an overnight culture. Thoroughly remove all medium.
2. Cell Suspension: Add 250ml of Cell Suspension Buffer (G1, containing RNase A), to the pellet and suspend the cell until homogeneous.
3. Cell Lysis: Add 250ml of Cell Lysis Solution (G2). Mix gently by inverting the capped tube five times. Do not vortex. Incubate at room temperature for 5 minutes.
4. Neutralization: Add 350ml of Neutralization Buffer (M3) and mix immediately by inverting the tube five times. Do not vortex. Centrifuge mixture at ? 12,000xg for 10 min.
5. Cartridge Loading: Place a cartridge in a 2ml wash tube. Load the supernatant from step 4 into the spin cartridge. Centrifuge mixture at ? 12,000xg for 1 min. Discard flow-through.
6. Cartridge Wash: Place the cartridge back into the 2ml wash tube. Add 700ml of Wash Buffer (G4, containing ethanol) to spin cartridge. Centrifuge mixture at ? 12,000xg for 1 min. Discard flow-through. Centrifuge again at ? 12,000xg for 1 min, to remove residual wash buffer.
7. Plasmid Elution: Place the cartridge into a 1.5ml recovery tube. Add 75ml of warm TE Buffer (65ºc-70ºc) directly to the center of the spin cartridge. Incubate at room temperature for 1 min, and then centrifuge mixture at ? 12,000xg for 2 min.