meth_3.1.10 MP Biomedicals FastDNA Spin Kit for Soil modified for soil crust and filtered water samples from acidic saline lakes

Contributor(s): Elizabeth Watkin (E.Watkin@curtin.edu.au); Katelyn Boase (katelyn.boase@postgrad.curtin.edu.au)

Citation:

AM Project ID: AM0019

Extractions performed using the MP Biomedicals FastDNA® Spin Kit for Soil (https://www.mpbio.com/116560000-fastdna-spin-kit-for-soil-samp-cf ), following the supplied protocol with the following alterations:

For filter papers: In 8 x separate extractions, use 1/8^th of pre-cut 90 mm filters (0.2 µm and 0.1 µm Nalgene™ Rapid-Flow™ Sterile Disposable Filter Units with PES membrane #catalogue numbers TBC) into the bead beating tubes*.

For sediment and salt crusts: Triplicates of lake samples were extracted and combined after elution (see below diagram). 0.5 g of sample was added into each bead beating tube.

NB. Skim control extraction performed with each extraction set to be sequenced.

• Step 1: ~40 mg.mL-1 steam sterilized skim milk powder was added to the Bead beating tubes prior to the addition of the samples.
• After step 3: five rounds of freeze thawing in liquid nitrogen and heating at 95 ℃.
• Step 4: three rounds of homogenisation at 6.5 m/s for 45 seconds in a Fast-Prep24TM ribolyser system with 5-minute rests on ice between cycles.

* TBC if using 1/8^th of the 0.2 µm and 0.1 µm filter paper into in a single extraction tube or, extract separately and combine the DNA for sequencing (this is to ensure capture of Nanoarchaea).