Sponsored,vendor-submitted protocol Sponsored by Mo Bio Laboratories, Inc.
Published in
November
2009
(p.19)
DOI: 10.2144/000113290
Abstract
Environmental researchers face the difficult task of studying microbial communities from remote locations and under extreme conditions. Two major issues facing soil microbiologists are the preservation of the original microbial population during transport and storage and the removal of enzyme inhibitors from DNA and RNA to ensure successful gene expression or genotyping analysis. Methods for the stabilization of soil samples during transport and options for the isolation of RNA or DNA from environmental samples are described.
Introduction
Environmental samples are inherently problematic due to inhibitors that cause PCR failure and inaccuracy in metagenomic analysis from irreplaceable samples. Typical inhibitors include humic acids, fulvic acids, polysaccharides, and polyphenols. Using the patented Inhibitor Removal Technology® (IRT), removal of PCR inhibitors from the final DNA or RNA is accomplished in even the most difficult samples. An equally important concern involves preservation of the microbial metatranscriptome in samples at the time of collection. A novel bacteriostatic solution that freezes the microbial content of soils during transport and storage and protects the RNA from degradation is the LifeGuard™ Soil Preservation Solution. Using LifeGuard™, soils stored at 4°C were stabilized, and total RNA was isolated suitable for terminal restriction fragment length polymorphism (T-RFLP) analysis. The combination of LifeGuard™ with the RNA PowerSoil® Kit results in a complete system for obtaining high quality RNA and DNA from soil.
Materials and methods
test LifeGuard™ Soil Preservation Solution, 100 mL (Cat. no. 12868-100; MO BIO Laboratories)
RNA PowerSoil® Total RNA Isolation Kit, 25 preps (Cat. no. 12866-25; MO BIO Laboratories)
RNA PowerSoil® DNA Elution Accessory Kit, 25 preps (Cat. no. 12867-25; MO BIO Laboratories)
PowerSoil® DNA Isolation Kit, 50 preps (Cat. no. 12888-50; Mo Bio Laboratories); 100 preps (Cat. no. 12888-100; MO BIO Laboratories)
Results
Preservation of microbial communities in Antarctic Dry Valley soil using LifeGuard™ Soil Preservation Solution
test T-RFLP results from LifeGuard™ stabilized soil indicate the soil microbial community was preserved during transport from Antarctica and New Zealand (Figure 1).
Isolation of high yields and quality RNA and DNA from the same soil sample using the DNA Accessory Kit
In Figure 2, the lanes are respresentative of the following: lane 1: ladder; lane 2: mulch RNA; lane 3: mulch DNA; lane 4: strawberry field RNA; lane 5: strawberry field DNA; lane 6: compost RNA; lane 7: compost DNA; lane 8: ladder.
The PowerSoil® DNA Isolation Kit gives 100% Success
PCR amplification of genomic DNA isolated from eight different soils using the PowerSoil®DNA Isolation Kit and kits from two other suppliers (Figure 3).
Only the MO BIO kit gives 100% success.
Summary
For preservation of soils and isolation of DNA and RNA from environmental samples, MO BIO offers the widest selection of methods suitable for metagenomic and metatranscriptome analysis. Trust irreplaceable samples to the experts at MO BIO Labs.
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