IGEM:British Columbia/Protocols/Miniprep

Materials

 * Overnight culture
 * Isopropanol
 * P1 buffer
 * P2 buffer
 * 3M sodium/potassiuma acetate
 * 70% ethanol
 * sdH2O

Equipment

 * Tabletop Centrifuge
 * Vortex
 * Water bath

Method

 * Spin down up to 5mL of overnight culture at max speed
 * Pipette out the broth; minimize broth as much as possible
 * Add 200uL of buffer P1 (50mM Tris-Cl pH 8.0, 10mM EDTA, 100ug/mL RNAse A), resuspend pellet in this buffer
 * Add 200uL of buffer P2 (200mM NaOH, 1%SDS), invert the tube 6-10 times and wait for 5 minutes at room temperature.
 * Add 300uL of 3M sodium/potassium acetate (pH 5.5), immediately invert the tube 6-10 times, and centrifuge for 10 minutes at 13,000 RPM.
 * Pour or pipette the supernatant into a microfuge tube with 600uL isopropanol, make sure to avoid the white precipitate as much as possible
 * Vortex the tube for 20 seconds and centrifuge for one minute at 13,000 RPM
 * Discard the supernatant and add 750uL of 70% ethanol to the tube, vortex briefly, and centrifuge for another 1 minute at 13,000 RPM
 * Discard the supernatant again and centrifuge for 10 seconds at 13,000 RPM to collect the remaining ethanol.
 * If you can see a pellet, pipette out the remaining ethanol, if not, heat the tube over a hot water bath (50C or more) for as long as it takes to evaporate the ethanol. A P10 tip works best because it won't freak out the ethanol as much when it comes in contact with the tip.
 * Resuspend the plasmid DNA in the 50µL of sterile distilled water.