An efficient procedure for the isolation of PCR-competent DNA from Bacillus endospores germinated in soil

DNA extraction techniques for endospore-forming bacteria in soil are often labour-intensive and unreliable. Our objective in this work was to investig ate whether good quality DNA could be obtained from spores germinated in so il. To this end, endospores from Bacillus subtilis, B. megaterium and B. th uringiensis were inoculated into soil microcosms and germination was induce d by addition of LB medium supplemented with l-alanine, glucose, fructose a nd KCl. Heat resistance count was reduced to 80% for B. subtilis and more t han 90% for B. thuringiensis and B. megaterium after a few minutes. Isolati on of DNA from soil with a procedure which did not work on spores was shown to be as efficient for in situ-germinated spores as for inoculated vegetat ive cells. Furthermore, we developed a simple procedure that allowed us to use the recovered DNA in PCR amplifications. The present methodology is sim ple and efficient; it avoids the use of special equipment and harsh spore r upturing methods and can be carried out with multiple samples.