min incubation at 70 °C, 200 μl ethanol (96%–100%) was then added and mixed again. The mixture was placed into the DNeasy Mini column and centrifuged at 6,000g (8,000 rpm) for 1 min. The DNeasy Mini spin column was then placed in a new 2-ml collection tube, and 500 μl buffer AW1 was added, followed by placement in a centrifuge for 1 min at 6,000g (8,000 rpm). The DNeasy Mini spin column was then placed in a new 2-ml collection tube again, and 500 μl buffer AW2 was added and centrifuged for 3 min at 20,000g (14,000 rpm) to dry the DNeasy membrane. Then the DNeasy Mini spin column was placed in a clean 1.5-ml or 2-ml microcentrifuge tube and 200 μl buffer AE was pipetted directly onto the DNeasy membrane. This was incubated at room temperature for 1 min and then centrifuged for 1 min at 6,000g (8,000 rpm) to elute. Two 200-μl elutions were performed followed by ethanol/sodium acetate precipitation and resuspension of the resultant pellet with TE buffer.
