Extraction, detection and persistence of extracellular DNA in forest litter microcosms

A DNA extraction method was developed that preferentially extracted extracellular DNA rather than intracellular DNA from forest litter. The method purposely avoided the use of harsh chemicals and physical disruption steps used in total DNA extraction to release DNA from cells. The detection limit of PCR, determined by spiking forest litter samples with a dilution series of Choristoneura fumiferana MNPVegt(-)/lacZ(+) genomic DNA, was about 1 ng DNA or 6.85 X 10(6) target copies 0.5 g(-1) moist forest litter or 0.14 g(-1) dry forest litter. In this study, outdoor terrestrial microcosms, each spiked with 49.2 mug of genomic DNA (from the baculovirus CJMNPVegt(-)/lacZ(+)), were exposed to summer conditions. A 530 by DNA fragment from the genome of the baculovirus CJMNPVegt(-)/lacZ(+) was detected in these microcosms for about 3 months. The DNA may have persisted for a longer period but was below the detection limit of the PCR analysis. (C) 2004 Elsevier Ltd. All rights reserved.