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, Close the lid and grind the samples using a Tissue Lyser II ® for 1 min at 30 Hz. Repeat once if necessary
, Add 63 µL of 20 % SDS (B) and 5 µL of Proteinase K (D)
, Vortex briefly 6. Incubate on Eppendorf Thermomixer ® at 55 °C and 900 rpm for 3 hours
, Transfer lysate into a 1.5 mL Safe Lock tube and retrieve the bead (it can be washed and reused)
, Add 400 µL of protein precipitation solution (C). Agitate by hand
, Add 600 µL of isopropanol at -20°C (E) and mix by turning the tubes upside down 30 times. 14. Incubate at -20°C overnight
, Discard the supernatant and keep the white pellet at the bottom of the tube. By pouring gently the supernatant into trash the pellet should stay stick to the tube
, Place the tubes upside up with open lid for 5 minutes at room temperature
, Close the lid and place the tubes at 50 °C for 10 minutes to resuspend DNA. Be sure to resuspend the DNA before proceeding with DNA purification
, Heat the DE buffer at 70 °C
, Add 525 µL od DB buffer into the 175 µL of resuspended DNA
, Place the NucleoSpin ® gDNA Clean-up Column on a collection tube and load the sample on the column
, Centrifuge at 11 000 g for30 seconds
, Add 700 µL of DW buffer to the column. Vortex for 2 seconds
, Add 700 µL of DW buffer to the column. Vortex for 2 seconds
, Add 50 µL of DE buffer (at 70 °C) on the column. Let it rest for 1 minute at room temperature with an open lid
, Add 50 µL of DE buffer (at 70 °C) on the column. Let it rest for 1 minute at room temperature with an open lid
, Stock DNA at -20°C