Sample preparation and lysis of homogenized malaise trap samples

Abstract

This protocol describes the steps of sample preparation and lysis before DNA extraction for the Malaise trap metabarcoding protocol of the LeeseLab.

Before start

Make sure all buffers are prepared before starting.

Steps

For each sample prepare a screwcap tube pre-filled with a few 2 mm zirconia beads.

Shake the sample well.

Transfer 800µL of the small size fraction and 200µL of the large size fraction to a 2 mL screwcap tube. It might be beneficial to use wide-bore tips or cut off the tip when using regular pipette tips.

11.000x g

Remove as much ethanol as possible with a 1000µL pipette.

Add 900µL of TNES buffer and 100µL of Proteinase K working solution . Vortex shortly.

Note

Depending on the amount of samples this can be prepared as a mastermix. We usually prepare TNES + Proteinase K in batches for 24 samples. Proteinase K tends to self-digest if the time for samples preparation takes too long.

Bead-beat for 0h 2m 0s at 2400rpm

Incubate 1400rpm

Store at -20°C until DNA extraction.

Abstract

Before start

Steps

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