DNA extraction from fungal mycelium using Extract - n - Amp
Turn heat block on and preheat to 95C .
Working at the lab bench , add ½ scoop of the 1.0 mm zirconium oxide beads to a sterile 1.5 ml tube .
Close the lid and label the top and side of each tube with the sample name ( e . g . , AK0013 ) .
In either the laminar flow hood or biosafety cabinet , remove a small piece ( no bigger than 0.5 cm2 ) of mycelium from the culture tube / plate and transfer to the corresponding labeled 1.5 ml tube .
Try to minimize the amount of agar .
* * * This step must be donin a sterile environment to ensure the cultures are not contaminated , but the remaining steps can be done on the lab bench .
Add 100 ul of Extraction Buffer ( making sure the mycelium is completely submerged in the liquid ; you may need to centrifuge ) and firmly close tube lids .
Place tubes in the bead beater .
If using the Bullet Blender Storm make sure the lid is for the 1.5 ml tubes not the screwcap tubes .
Close the lid , and bead beat for 1 minute on speed 10 .
( Alternatively you can grind the tissue with a sterile blue pestle . )
* * Use ear protection when using the bead beater .
Briefly ( < 30 sec ) centrifuge to remove liquid from the lid of the tube .
* * * Do not leave samples in the Extraction Buffer > 25 minutes before proceeding to the heating step .
Place tubes in heat block at 95°C for 10 minutes ; then briefly centrifuge after incubation to remove condensation .
Add 100 ul of Dilution Buffer and vortex to mix .
Store the DNA at 2 - 8°C for short - term use and - 20° to - 80° for long - term storage .
