BIOLOGY, BIOLOGICAL CONTROL, AND MOLECULAR GENETICS OF ROOT DISEASES OF WHEAT, BARLEY AND BIOFUELS BRASSICAS
Location: Root Disease and Biological Control Research
Title: Improved quantification of pathogen DNA from soil using pressure cycling technology
Submitted to: International Congress of Plant Pathology Abstracts and Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: June 20, 2008
Publication Date: August 20, 2008
Citation: Okubara, P.A., Schroeder, K.L., Li, C., Schumacher, R.T., Lawrence, N.P. 2008. Improved quantification of pathogen DNA from soil using pressure cycling technology. International Congress of Plant Pathology Abstracts and Proceedings.
Detection and quantification of Rhizoctonia, Pythium and other soilborne pathogens are inconsistent at low pathogen populations and in hard-to-extract samples, despite use of sensitive diagnostic assays such as real-time PCR. An efficient and reproducible extraction system in which samples are subjected to cycles of hydrostatic pressure (from ambient to as high as 35,000 psi) under controlled conditions has been used to improve the extraction of DNA from Rhizoctonia and Pythium in soil samples. This novel technology, called Pressure Cycling Technology (PCT SPS, Pressure BioSciences, Inc., West Bridgewater, Massachusetts, USA) improved the extraction of Rhizoctonia and Pythium DNA from agricultural soils up to 30-fold and 2-fold, respectively, compared to samples without the pressure cycling treatment. Using the PCT SPS, we obtained detectable amounts of Rhizoctonia DNA from wheat roots that were previously recalcitrant to homogenization. In 80% of the root samples, pathogen DNA was extracted in amounts high enough to be quantified using real-time PCR. Furthermore, as the PCT SPS is a closed system, samples were free from contamination that can occur during mortar-and-pestle extractions. The PCT SPS confers a significant advantage in germplasm screening, food security assessment, and characterization of host and microbe nucleic acids.