About ZR-96 Fecal DNA Kit™
The ZR-96 Fecal DNA Kit™ is designed for the simple and rapid isolation of inhibitor-free, PCR-quality host cell and microbial DNA from a variety of sample sources including humans, birds, rats, mice, cattle, etc. The procedure is easy and can be completed in minutes: fecal samples are rapidly and efficiently lysed by bead beating with our state of the art, ultra-high density BashingBeads™. Fast-Spin plate technology is then used to isolate the DNA which is subsequently filtered to remove humic acids/polyphenols that can inhibit PCR. Eluted DNA is ideal for downstream molecular-based applications including PCR, arrays, genotyping, methylation detection, etc.
|DNA Recovery||Typically, up to 5 µg total DNA is eluted into ≥ 25 µl DNA Elution Buffer per sample.|
|Processing Time||50 min|
|Equipment||Centrifuge w/ microplate carriers, 96-well plate/block disruptor or pulverizer.|
|DNA Size Limits||Capable of recovering genomic DNA sized fragments from 100 bp to ≥40 kb. Typical fragment sizes range from 25 kb "“ 35 kb. If present, parasitic and viral DNA will also be recovered.|
|Sample Sources||Bacterial, protist, as well as host DNA can be effectively isolated from a ≤80 mg sample of mammalian feces.|
|DNA Purity||High quality, inhibitor-free DNA is eluted with DNA Elution Buffer suitable for the amplification of bacterial, protist, and/or mammalian templates. Absorbance 260 nm/280 nm >1.8.|
Comparison of DNA yields from rat feces using the ZR Fecal DNA MiniPrep™ and kits from suppliers A and B. Equivalent amounts of feces were processed using each kit and then equal volumes of eluted DNA were analyzed in a 0.8% (w/v) agarose/ethidium bromide gel. Samples were processed in triplicate.
Add up to 80 milligrams of fecal sample to the tubes of a ZR BashingBead™ Lysis Rack. Add 400 µl Lysis Solution to each tube. Cap tubes tightly to prevent leakage.
Secure in a 96-well block/plate bead beater (e.g., 2010 GenoGrinder®) and process samples. Optimization of processing time/speed will be necessary for complete sample lysis.
Processing times may be as little as one minute when using high-speed cell disrupters (e.g., 2000 GenoGrinder®, page 5). See manufacturer’s literature for operating information.
Centrifuge the ZR BashingBead™ Lysis Rack at ≥ 3,000 x g (5,000 x g max.) for 5 minutes.
Transfer up to 250 µl supernatant to each well of a Deep-Well Block.
Add 750 µl of Fecal DNA Binding Buffer to the supernatant in the Deep-Well Block from Step 4. Cover completely with Cover Foil and mix thoroughly by vortexing for 2 minutes. Centrifuge the Deep-Well Block at ≥ 3,000 x g (5,000 x g max.) for 5 minutes.
Remove or pierce foil and transfer 500 µl of each of the mixtures from Step 5 to the wells of a Silicon-A™ Plate on a Collection Plate. Centrifuge the assembly at ≥ 3,000 x g (5,000 x g max.) for 5 minutes.
Discard the flow through from the Collection Plate and repeat Step 6.
Add 200 µl DNA Pre-Wash Buffer to the wells of the Silicon-A™ Plate on the emptied Collection Plate and centrifuge the assembly at ≥ 3,000 x g for 5 minutes.
Add 500 µl Fecal DNA Wash Buffer to the wells of the Silicon-A™ Plate on the Collection Plate and centrifuge the assembly at ≥ 3,000 x g for 5 minutes.
Transfer the Silicon-A™ Plate onto a clean Elution Plate and add 100 µl (50 µl minimum) DNA Elution Buffer directly to the matrices in the plate. Centrifuge the assembly at ≥ 3,000 x g for 5 minutes.
Transfer the eluted DNA from Step 10 to a prepared Silicon-A™-HRC Plate mounted onto a clean Elution Plate. Centrifuge the assembly at exactly 3,500 x g for 3 minutes.
Eluted, ultra-pure DNA is now ready for use in your experiments, or the Elution Plate can be covered with Cover Foil for storage of the DNA.