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"Language": "en",
"Value": "Store the kit at +15 to +25°C.",
"Country": "XG",
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Solution | Storage |
DNase solution | -15 to -25°C |
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"Value": "For small-scale (mini) preparations of RNA",
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Vial / Bottle | Cap | Label | Function / Description | Content |
---|
1 | green | Lysis/-Binding Buffer | 4.5 M guanidine-HCl, 50 mM Tris-HCl, 30% Triton X-100 (w/v), pH 6.6 (25°C) | 25 ml |
2 | | DNase I, recombinant, lyophilizate | 10 KU lyophilized DNase I | Resuspend in 0.55 ml Elution Buffer |
3 | white | DNase Incubation Buffer | 1 M NaCl, 20 mM Tris-HCl and 10 mM MnCl2, pH 7.0 (25°C) | 10 ml |
4 | black | Wash Buffer I | 5 M guanidine hydrochloride and 20 mM Tris-HCl, pH 6.6 (25° C); final concentrations after addition of 20 ml absolute ethanol | 33 ml (add 20 ml absolute ethanol before first use) |
5 | blue | Wash Buffer II | 20 mM NaCl, 2 mM Tris-HCl, pH 7.5 (25°C); final concentrations after addition of 40 ml absolute ethanol | 10 ml (add 40 ml absolute ethanol before first use) |
6 | colorless | Elution Buffer | Water, PCR Grade | 30 ml |
7 | | High Pure Filter Tubes | | One bag with 50 polypropylene tubes with two layers of glass fiber fleece, for uptake of up to 700 μl sample volume |
8 | | Collection Tubes | | One bag with 50 polypropylene tubes (2 ml) |
All solutions are clear and should not be used when precipitates have formed. Warm the solutions at +15 to +25°C or in a 37°C water bath until the precipitates have dissolved.
The buffers can show a slight yellow color. This will have no impact on the function of the buffer.
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"Code": "Content",
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"Value": "Isolation of RNA is a prerequisite for the analysis of gene expression. Frequently applied techniques like reverse transcriptase-PCR (RT-PCR), northern blotting, RNase protection and primer extension require the use of intact, undegraded RNA from different sample materials like cultured cells, blood, yeast and bacteria.
Samples are lysed and homogenized in the presence of chaotropic salts, then applied to the spin filter tube. Nucleic acids bind specifically to the surface of the filter. Co-purified DNA is ultimately digested with DNase I. The bound RNA is purified from salts, proteins, digested DNA, and other impurities by washing steps, followed by an elution.
- Cultured cells are lysed by a special Lysis/-Binding buffer. At the same time, RNases are inactivated.
Other sample materials require a specific pre-lysis treatment.
- Nucleic acids are bound to the glass fibers pre-packed in the High Pure Filter Tube.
- Residual contaminating DNA is digested by DNase I, applied directly onto the glass fiber fleece.
- Bound nucleic acids are washed with a special Inhibitor Removal Buffer to get rid of RT-PCR inhibitory contaminants.
- Further washing of bound nucleic acids purifies them from salts, proteins, and other cellular impurities.
- RNA is recovered using the elution buffer.
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"Value": "The High Pure RNA Isolation Kit is designed for the purification of total RNA from cultured cells. Other sample materials like blood, yeast and bacteria require an additional specific pre-lysis treatment, which is described in the protocol section.
Due to the integrated DNase digestion step, contamination of the isolated RNA with residual genomic DNA is mostly avoided. In addition, RNA is suited for other techniques like northern blotting, RNase protection and primer extension. Up to 24 samples can be processed simultaneously in approx. 1 hour. Thus, the purification procedure is less time consuming compared with alternative methods which require extraction with organic solutions, RNA precipitation or ultracentrifugation.",
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Total time | Approximately 1 hour (24 samples simultaneously) |
---|
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