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Extraction of High Molecular Weight Genomic DNA from Soils and Sediments
Published on: November 10, 2009
Flotation as a tool for indirect DNA extraction from soil.
Nádia Skorupa Parachin1, Jenny Schelin, Börje Norling
1Applied Microbiology, Lund Institute of Technology, Lund University, PO Box 124, 221 00 Lund, Sweden.
Applied Microbiology and Biotechnology
|June 10, 2010
Summary
This study introduces a new soil DNA extraction method using microbial cell flotation. It efficiently isolates pure, high-yield environmental DNA, overcoming inhibition issues for molecular analyses.
Area of Science:
- Environmental Science
- Molecular Biology
- Microbiology
Background:
- Soil microbial diversity is assessed using total DNA extraction.
- Co-extraction of inhibitory substances hinders downstream molecular applications like PCR.
- Improved DNA extraction methods are needed for pure DNA with satisfactory yield.
Purpose of the Study:
- To develop a novel soil sample preparation protocol for efficient environmental DNA (eDNA) extraction.
- To separate microbial cells from soil matrix and inhibitory substances using flotation.
- To achieve high DNA yield and purity suitable for molecular analyses.
Main Methods:
- Utilized buoyant densities of soil microorganisms for cell separation.
- Designed a discontinuous density gradient with silanised silica particles (BactXtractor).
- Employed a flotation-based separation technique to isolate microbial cells.
Main Results:
- The flotation method efficiently separated microbial cells from the soil matrix.
- Extracted eDNA exhibited high molecular weight and comparable yields (3.5 µg/g soil) to commercial kits.
- Extracted DNA was free of inhibitors, allowing successful PCR and restriction enzyme digestion.
- Both prokaryotic and eukaryotic sequences were recovered using specific primers.
Conclusions:
- The developed flotation method is an efficient alternative to direct soil DNA extraction protocols.
- This technique yields pure, high-quality eDNA suitable for diverse molecular applications.
- The method effectively removes enzyme-inhibitory substances present in soil samples.
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