Use of pressure cycling technology for cell lysis and recovery of bacterial and fungal communities from soil

Emily A Bruner; Patricia A Okubara; Rita Abi-Ghanem; David J Brown; Catherine L Reardon

doi:10.2144/000114273

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Use of pressure cycling technology for cell lysis and recovery of bacterial and fungal communities from soil

Journal article

Open access

Peer reviewed

Use of pressure cycling technology for cell lysis and recovery of bacterial and fungal communities from soil

Emily A Bruner, Patricia A Okubara, Rita Abi-Ghanem, David J Brown and Catherine L Reardon

BioTechniques, Vol.58(4), pp.171-180

04/2015

DOI: https://doi.org/10.2144/000114273

Handle:

https://hdl.handle.net/2376/109611

PMID: 25861929

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Abstract

DNA, Bacterial - isolation & purification

Polymorphism, Restriction Fragment Length

Microbiota - genetics

Soil Microbiology

DNA, Bacterial - chemistry

DNA, Fungal - chemistry

DNA, Fungal - isolation & purification

Hydrostatic Pressure

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.

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Title: Use of pressure cycling technology for cell lysis and recovery of bacterial and fungal communities from soil
Creators: Emily A Bruner - Department of Crop and Soil Sciences, Washington State University, Pullman, WA
Patricia A Okubara - Root Disease and Biological Control Research, USDA-ARS, Pullman, WA
Rita Abi-Ghanem - Department of Crop and Soil Sciences, Washington State University, Pullman, WA
David J Brown - Department of Crop and Soil Sciences, Washington State University, Pullman, WA
Catherine L Reardon - Columbia Plateau Conservation Research Center, USDA-ARS, Pendleton, OR
Publication Details: BioTechniques, Vol.58(4), pp.171-180
Academic Unit: Crop and Soil Sciences, Department of; Plant Pathology, Department of
Publisher: England
Identifiers: 99900546920001842
Language: English
Resource Type: Journal article