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. 2020 May 20;8(5):763. doi: 10.3390/microorganisms8050763

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© 2020 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

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Procedure for efficient extraction of DNA from within B. anthracis spores. Individual experimental steps and respective time frames are indicated: (1) 1 mL of a spore suspension is added to 200 mg of PowerBead Tubes (0.1 mm); (2) tubes are processed for 5 min at 30 Hz in a TissueLyser II instrument; (3) the spore suspension is centrifuged for 10 min at 13,100× g; (4) the supernatant is concentrated using Microcon centrifugal filters to a final volume of 150 µL; (5) the concentrate is used for DNA extraction with the MasterPure kit (standard manufacturer’s instructions for fluid samples); (6) additionally, the debris pellet from the centrifugation step (3) is used for DNA extraction with the modified (see text for details) MasterPure protocol; (7) DNA extracted from the debris pellet and DNA isolated from lysis supernatant are each dissolved in 25 µL TE buffer and pooled.