A lipid extraction and analysis method for characterizing soil microbes in experiments with many samples

Lawrence G. Oates, Harry W. Read, Jessica L.M. Gutknecht, David S. Duncan, Teri B. Balser, Randall D. Jackson

Research output: Contribution to journalArticlepeer-review

12 Scopus citations

Abstract

Microbial communities are important drivers and regulators of ecosystem processes. To understand how management of ecosystems may affect microbial communities, a relatively precise but effort-intensive technique to assay microbial community composition is phospholipid fatty acid (PLFA) analysis. PLFA was developed to analyze phospholipid biomarkers, which can be used as indicators of microbial biomass and the composition of broad functional groups of fungi and bacteria. It has commonly been used to compare soils under alternative plant communities, ecology, and management regimes. The PLFA method has been shown to be sensitive to detecting shifts in microbial community composition. An alternative method, fatty acid methyl ester extraction and analysis (MIDI-FA) was developed for rapid extraction of total lipids, without separation of the phospholipid fraction, from pure cultures as a microbial identification technique. This method is rapid but is less suited for soil samples because it lacks an initial step separating soil particles and begins instead with a saponification reaction that likely produces artifacts from the background organic matter in the soil. This article describes a method that increases throughput while balancing effort and accuracy for extraction of lipids from the cell membranes of microorganisms for use in characterizing both total lipids and the relative abundance of indicator lipids to determine soil microbial community structure in studies with many samples. The method combines the accuracy achieved through PLFA profiling by extracting and concentrating soil lipids as a first step, and a reduction in effort by saponifying the organic material extracted and processing with the MIDI-FA method as a second step.

Original languageEnglish (US)
Article numbere55310
JournalJournal of Visualized Experiments
Volume2017
Issue number125
DOIs
StatePublished - Jul 16 2017

Bibliographical note

Funding Information:
This work was funded in part by the DOE Great Lakes Bioenergy Research Center (DOE BER Office of Science DE-FC02-07ER64494) and DOE OBP Office of Energy Efficiency and Renewable Energy (DE-AC05-76RL01830). The authors would like to acknowledge Anders Gurda for his patient and skillful contribution to videography and editing.

Publisher Copyright:
© 2017, Journal of Visualized Experiments. All rights reserved.

Keywords

  • Bacteria
  • Environmental sciences
  • Fatty acid methyl ester (FAME)
  • Fungi
  • Issue 125
  • Lipid biomarker
  • MIDI-FA
  • Microbial community
  • Ordination
  • Phospholipid fatty acid (PLFA)
  • Soil microbiology

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