Kasson Lab DNA Extraction

Turn on hot water bath, set to 65°C.

Pull two Eppendorf 1.5mL centrifuge tubes per sample.

Label both sets of tubes with (short) sample names.

Label one tube set for each sample with an "I" for .

<img src="https://static.yanyin.tech/literature_test/protocol_io_true/protocols.io.q26g78413lwz/Screen%20Shot%202020-10-28%20at%201.12.23%20PM.png" alt="Sketch of "I"-labeled tubes (Angie Macias)." loading="lazy" title="Sketch of "I"-labeled tubes (Angie Macias)."/>

Add 600µL of (or ) to tubes without "I" .

Add 600µL of to tubes labeled with "I" .

Place tube with into 65°C water bath.

Sterilize some metal scrapers with flame and 95% (v/v) .

Add 1/2 pea-sized amount of fungal tissue (young hyphae) to each tube containing .

Flame-sterilize and cool scrapers between samples.

Macerate each sample with a new, sterile micropestle until tissue is homogenous.

Add tubes to a floating rack to allow samples to incubate directly in 65°C water bath for 0h 30m 0s.

Remove samples and vortex for 0h 0m 3s before returning to 65°C water bath for 0h 30m 0s.

Place a sufficient aliquot of in water bath to warm for Step 21.

Remove samples and allow them to cool on the bench for 0h 5m 0s.

Add 200µL of to each tube and vortex for 10 seconds.

Centrifuge samples for 0h 3m 0s at 14.000rpm,0h 0m 0s.

Using a P1000 micropipette, transfer supernatant to each tube containing and gently mix by inversion.

Centrifuge for 0h 1m 0s at 14.000rpm,0h 0m 0s.

Carefully pour off the supernatant into waste container.

Add 600µL of 70% (v/v) to each tube and mix gently by inversion.

Centrifuge for 0h 1m 0s at 14.000rpm,0h 0m 0s.

Repeat Step 16.

Open and invert tubes onto a clean paper towel.

Add 100µL of warmed to each tube.

Store fully-labeled tubes in a box (not a tube rack) in the -20°C freezer.