title: Use of pressure cycling technology for cell lysis and recovery of bacterial and fungal communities from soil.
abstract: Selection of cell lysis methodology is critical to microbial community analyses due to the inability of any single extraction technology to recover the absolute genetic structure from environmental samples. Numerous methodologies are currently applied to interrogate soil communities, each with its own inherent bias. Here we compared the efficacy and bias of three physical cell lysis methods in conjunction with the PowerLyzer PowerSoil DNA Isolation Kit (MO BIO) for direct DNA extraction from soil: bead-beating, vortex disruption, and hydrostatic pressure cycling technology (PCT). PCT lysis, which is relatively new to soil DNA extraction, was optimized for soils of two different textures prior to comparison with traditional bead-beating and vortex disruption lysis. All cell lysis methods successfully recovered DNA. Although the two traditional mechanical lysis methods yielded greater genomic, bacterial, and fungal DNA per gram soil than the PCT method, the latter resulted in a greater number of unique terminal restriction fragments by terminal RFLP (T-RFLP) analysis. These findings indicate the importance of diversity and quantity measures when assessing DNA extraction bias, as soil DNA retrieved by PCT lysis represented populations not found using traditional mechanical lysis methods.
authors: Department of Crop and Soil Sciences, Washington State University, Pullman, WA.
authors: Emily A
authors: EA
authors: Bruner
authors: Root Disease and Biological Control Research, USDA-ARS, Pullman, WA.
authors: Patricia A
authors: PA
authors: Okubara
authors: Department of Crop and Soil Sciences, Washington State University, Pullman, WA.
authors: Rita
authors: R
authors: Abi-Ghanem
authors: Department of Crop and Soil Sciences, Washington State University, Pullman, WA.
authors: David J
authors: DJ
authors: Brown
authors: Columbia Plateau Conservation Research Center, USDA-ARS, Pendleton, OR.
authors: Catherine L
authors: CL
authors: Reardon
mesh_terms: D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification*; D004271:DNA, Fungal / Q000737:chemistry / Q000302:isolation & purification*; D006874:Hydrostatic Pressure; D064307:Microbiota / Q000235:genetics*; D012150:Polymorphism, Restriction Fragment Length; D012988:Soil Microbiology
chemical_list: D004269:DNA, Bacterial; D004271:DNA, Fungal
keywords: DNA extraction; T-RFLP; lysis method; microbial community; pressure cycling technology (PCT); quantitative PCR (qPCR); soil