Principle

Speedy isolation of total RNA from various stool specimen

The NucleoSpin® RNA Stool kit is designed for the efficient isolation of RNA including small RNA species from fresh and frozen stool samples.
The kit combines NucleoZol and a specific Lysis Buffer RST1 for a chemical disruption of the stool sample and the containing microbes. The mechanical lysis is performed using NucleoSpin® Bead Tubes Type A (containing ceramic beads) in combination with a mechanical disruption device. No enzymatic reactions like protease digestion are required to homogenize the sample material. Undissolved sample material and the ceramic beads are subsequently removed by a short centrifugation. A next lysate clearing step comprises addition of Binding Buffer RST2, followed by a short centrifugation. The supernatant is finally cleared by passing it through a NucleoSpin® Inhibitor Removal Column that removes substances in stool samples that interfere with qRT-PCR. Binding conditions are adjusted by adding additional Binding Buffer RST2 to the flowthrough of the NucleoSpin® Inhibitor Removal Column. Next the sample is loaded onto a NucleoSpin® RNA Stool Column.
Residual contaminants such as complex polysaccharides, bile salts, and other PCR inhibitors are removed by an efficient washing procedure using Buffer RST2 as well as Wash Buffers RST3, RST4 and RST5. In between the different washing steps, residual DNA is removed via an on column DNA digestion step using rDNase. After a drying step, ready-to-use RNA can be eluted with RNase free H2O.

NucleoSpin® RNA Stool procedure

Features

• Combination of mechanical homogenization and chemical lysis ensures optimized RNA yield, including small RNA
• Isolated RNA does not show any PCR inhibition due to NucleoSpin® Inhibitor Removal Columns
• Suitable for various human and animal stool samples

Applications*

• RNA isolation from stool samples
• Typical downstream applications: real-time RT-PCR, Northern blotting, primer extension, array technology, RNase protection assays

* Kits to be used for research purposes only