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- Title
Extraction and detection of baculoviral DNA from lake water, detritus and forest litter.
- Authors
England; Trevors; Holmes
- Abstract
Aims: This paper describes a quick, reproducible, sensitive method for baculoviral DNA extraction, purification and detection from freshwater and forest litter environments. Methods and Results: The extraction protocol utilizes enzymatic and chemical lysis and physical disruption. To assess the efficiency of the extraction and purification protocol, PCR was used to detect a 530 bp DNA fragment from the genome of a genetically-modified baculovirus, Choristoneura fumiferana NPVegt–/lacZ+. The detection limit of PCR amplification was routinely about 4·1 × 102 occlusion bodies (OBs) 450 μl–1 lake water. Template DNA from the detritus and forest litter samples required 100-fold dilutions before use in PCR reactions. The detection limits for detritus and forest litter samples were routinely about 7·41 × 103 and 2·08 × 104 OBs 0·5 g–1 dry weight, respectively. Conclusions: The DNA extraction and purification methodology is reproducible, sensitive and can be used in lieu of, or in conjunction with, insect bioassays. Significance and Impact of the Study: The DNA extraction and purification protocol described in this paper will facilitate risk assessment and ecological studies of both wild-type and genetically-modified baculoviruses.
- Subjects
BACULOVIRUS genetics; DNA; DETRITUS; FOREST litter
- Publication
Journal of Applied Microbiology, 2001, Vol 90, Issue 4, p630
- ISSN
1364-5072
- Publication type
Article
- DOI
10.1046/j.1365-2672.2001.01289.x