About ZR Soil Microbe DNA MiniPrep™
The ZR Soil Microbe DNA MiniPrep™ is designed for the simple and rapid isolation of humic-free, PCR-quality DNA from microbes in soil. This product can be used to isolate DNA from tough-to-lyse bacteria, fungi, protozoa, and algae that inhabit a variety of samples including clay, sandy, silty, peaty, chalky, and loamy soils. Soil microbes are rapidly and efficiently lysed by bead beating with our state of the art, ultra-high density BashingBeads™. Fast-Spin column technology is then used to isolate the DNA, which is subsequently filtered to remove humic acids/polyphenols that can inhibit PCR. The procedure can be performed in minutes, and there is no need for organic denaturants or proteinases.
|DNA Recovery||Typically, up to 25 µg total DNA is eluted into ≥ 25 µl DNA Elution Buffer per sample.|
|Processing Time||15 min|
|Equipment||Microcentrifuge, vortex, cell disruptor/pulverizer (optional).|
|DNA Size Limits||Capable of recovering genomic DNA up to and above 40 kb. In most instances, mitochondrial DNA and viral DNA (if present) will also be recovered.|
|Sample Sources||DNA is isolated from bacteria, fungi, protozoa, and algae in up to 0.25 g of soil or from fungal/bacterial cells directly.|
|DNA Purity||High-quality, humic-free DNA is eluted with DNA Elution Buffer making it perfect for PCR. A260/A280 >1.8|
Metagenomic DNA isolated from 5 soil samples. M: 1 kb marker (NEB); 1-5: soil samples (sand, sandy clay loam, hydrophobic sandy loam, course sandy loam, fine gravel).
Add up to 0.25 grams of soil sample to a ZR BashingBead™ Lysis Tube. Add 750 µl Lysis Solution to the tube.
Alternatively, add 50-100 mg (wet weight) fungal and/or bacterial cells that have been resuspended in up to 200 µl of water or isotonic buffer (e.g., PBS) to a ZR BashingBead™ Lysis Tube.
Secure in a bead beater fitted with a 2 ml tube holder assembly (e.g., Disruptor Genie™) and process at maximum speed for 5 minutes.
Processing times may be as little as 40 seconds when using high-speed cell disrupters (e.g., the portable Xpedition™ Sample Processor, page 6, FastPrepâ-24, or similar). See manufacturer’s literature for operating information.
Centrifuge the ZR BashingBead™ Lysis Tube in a microcentrifuge at 10,000 x g for 1 minute.
Transfer up to 400 µl supernatant to a Zymo-Spin™ IV Spin Filter (orange top) in a Collection Tube and centrifuge at 7,000 x g for 1 minute.
Add 1,200 µl of Soil DNA Binding Buffer to the filtrate in the Collection Tube from Step 4.
Transfer 800 µl of the mixture from Step 5 to a Zymo-Spin™ IIC Column in a Collection Tube and centrifuge at 10,000 x g for 1 minute.
Discard the flow through from the Collection Tube and repeat Step 6.
Add 200 µl DNA Pre-Wash Buffer to the Zymo-Spin™ IIC Column in a new Collection Tube and centrifuge at 10,000 x g for 1 minute.
Add 500 µl Soil DNA Wash Buffer to the Zymo-Spin™ IIC Column and centrifuge at 10,000 x g for 1 minute.
Transfer the Zymo-Spin™ IIC Column to a clean 1.5 ml microcentrifuge tube and add 100 µl (25 µl minimum) DNA Elution Buffer directly to the column matrix. Centrifuge at 10,000 x g for 30 seconds to elute the DNA.
If fungi or bacterial cultures were sampled, the DNA is now suitable for PCR as well as other downstream applications.
Transfer the eluted DNA from Step 10 to a prepared Zymo-Spin™ IV-HRC Spin Filter (green top) (see above) in a clean 1.5 ml microcentrifuge tube and centrifuge at exactly 8,000 x g for 1 minute. The filtered DNA is now suitable for PCR and other downstream applications.
Archaeal and bacterial DNA was isolated from hypersaline water membrane filter samples using the ZR Soil Microbe DNA MiniPrep™. The DNA was then amplified by qPCR and followed by sequencing. Analysis of SSU rRNA gene amplicons revealed diverse prokaryotic communities with physiological functions involving carbon degradation and sulfate reduction.
The ZR Soil Microbe DNA MiniPrep™ was used to isolate microbial DNA from composted food waste and the resulting clone libraries containing 16s rRNA eubacteria and actinobacteria genes were used to identify the microbial community and characterize functional genes responsible for biotransformation.