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Nucleic acid purification

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NucleoSpin 96 DNA Stool product page
Homogenization bead selection guide
Product overview

High-throughput DNA purification from stool

With the refinement and broader application of next-generation sequencing technologies, researchers in the fields of medical, veterinary, and nutritional sciences have increasingly turned to genomics for meaningful answers regarding the role of gut microbiota in health and disease. While silica-membrane purification technology is well established as the method of choice for isolating high-quality DNA and efficiently removing inhibitors from animal and human fecal samples, this approach can prove challenging in a high-throughput context where columns may be subject to clogging during vacuum processing. An additional challenge involves the ever-present danger of contamination with humic acids, which can be a detriment to downstream analysis.

The NucleoSpin 96 DNA Stool kit has been designed to help researchers overcome these challenges by providing a reliable, automation-friendly method for DNA isolation—combining a robust filtration step to greatly reduce the likelihood of column-clogging with an efficient DNA extraction procedure based on silica-membrane technology. Provided below are application data demonstrating automated processing of stool samples on the epMotion 5075v platform. Processing of 96 samples using this approach takes ~100 minutes to complete.

At-a-glance  

Technology Silica-membrane technology combined with MN Bead Tubes Type A
Starting material 100–200 mg of human stool* (fresh or frozen)
Format 96-well plate
Handling Vacuum (manual or automated)
Fragment size 50 bp–approximately 50 kb
Typical yield 3–15 µg (varies by sample and disruption device)
A260/A280
 1.6–1.8
Elution volume 100–200 µl
Preparation time 90 min/96 preps (automated processing)
Binding capacity 50 µg


*For animal stool specimens and challenging samples with high amounts of lipids, polysaccharides, or proteins, it is recommended to first attempt processing of smaller inputs (e.g., 50–100 mg of sample material).

Manual vs. automated processing  

Comparable performance with manual or automated processing using NucleoSpin 96 DNA Stool.

Comparable performance with manual or automated processing. Six different stool samples were processed either manually or on an epMotion 5075v using the NucleoSpin 96 DNA Stool kit. Resulting DNA yields (blue and gray bars) and purity (orange dots) were comparable between manual and automated workflows for each sample.

DNA quality  

High-quality DNA from rat stool samples processed using NucleoSpin 96 DNA Stool.

High-quality DNA from rat stool samples. Eight different rat stool samples were processed using NucleoSpin 96 DNA Stool. Analysis of purity and yield of DNA isolated from each sample confirmed the reliably high quality of the extracted DNA.

Removal of inhibitors  

Efficient removal of PCR inhibitors with NucleoSpin 96 DNA Stool.

Efficient removal of PCR inhibitors with NucleoSpin 96 DNA Stool. DNA was purified from a human stool sample and sequentially diluted 1:4 up to a 4096-fold dilution. Diluted DNA and a reference standard were analyzed by qPCR using E. coli-specific primers and compared on a logarithmic plot. The slope of the resulting regression curve (–3.4106) indicated excellent qPCR performance without inhibition across the dilution series.

Downstream performance  

Consistently excellent PCR performance with NucleoSpin 96 DNA Stool.

Consistently excellent PCR performance. Six different human stool samples were processed either manually or on an epMotion 5075v using the NucleoSpin 96 DNA Stool kit in quadruplicates. 2.5-μl inputs from each eluate were used in a 25-μl endpoint PCR reaction designed to amplify a 1.5-kb portion of the bacterial 16S rRNA gene, and 3-µl outputs from each reaction were analyzed on a 1% agarose gel. Samples were successfully amplified in each case, demonstrating the robust downstream performance of DNA isolated using the kit.

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