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MagSi-DNA allround beads reversibly bind DNA and other nucleic acids under sample- and buffer-specific conditions. They can be used to isolate and purify nucleic acids from various (complex) mixtures, such as cell lysates, or enzymatic reactions. It works as following:A solution containing DNA (e.g. lysate) is combined with MagSi-DNA allround beads and a application-specific binding buffer. After a short incubation, nucleic acids are bound to the silica surface. By applying a suitable magnet to the container (tube/deep-well microplate) the bead pellet is separated from the sample mixture.Unwanted components are further removed by 3-4 washing steps in a selection of buffers (alcohol/water solutions). Nucleic acids are released in Dnase/RNase-free water or buffer solution (e.g. Tris, Tris-EDTA, pH 8).Silica and carboxyl (COOH) surfaces, but also nucleic acids, are negatively charged at neutral or basic pH, while both are also hydrated. For the binding mechanism of DNA to particles, dehydration is needed. This can be achieved by for instance alcohol, or by chaotropic agents such as guanidinium salts. Negative charges on the bead surface and the nucleic acid backbones are bridged by divalent cations. This can be reversed by a water solution. For washing, mostly alcohol/water mixtures are used, which will keep the DNA in dehydrated form and bound to the beads. To reduce premature elution of DNA, salts can be added to the washing solution. Elution takes place in a low-salt non-alcohol condition.
MagSi-DNA (COOH) beads are suspended in filtered water. The beads should be pre-washed in sterile filtered water to avoid any impact in downstream applications. The suspension media can be replaced with your own buffer/storage media. The beads are compatible with typical organic solvents like ethanol or isopropanol. However, chemicals with strong redox-portential should be avoided. The beads are stable in a pH range from 3 to 13 and at temperatures up to 95 degrees. After extensive incubations in these conditions, no degradation is measurable using spectrophotometric assays. Nevertheless, if you expect any interference in downstream applications, we strongly advise you to rinse the beads before use.
Lyse your cell, tissue, or bacterial sample via: mechanical disruption (sonication/French press) Enzymatic (lysozyme) methods Using a surfactant like Tween 20/SDS/Triton X-100. Lysing may be improved by heating the sample mixture.
Binding Add the binding buffer of choice to the lysate and mix well to get a homogeneous suspension. Add beads. Mix beads by vortexing before adding them to the sample. Depending on the expected amount of DNA the volume of beads can be varied. A good starting point is 20 μL when having 400-800 μL of cell lysate. Mix sample and incubate 2-10 minutes to allow the DNA to bind to the bead surface.Washing Following incubation, place the sample tube in a magnetic separator. In order to collect the beads to the side of the tube. Wait 30 seconds until all the beads have been attracted to the magnet. Discard the supernatant using a pipette, then remove the tube from the separator. Add 400 μL wash buffer, vortex 10 seconds and place the sample tube in a magnetic separator in order to collect the beads and discard the supernatant. Wash the beads at least twice.ElutionThe Elution buffer consists of a nuclease-free, non-alcohol solution (TE-buffer) to rehydrate the DNA so it will elute from the bead. Concentrated TE-buffer can be added to the pure sample to improve storage properties. Elute DNA by adding 50-200 μL elution buffer, pre-heated to 56 °C. Incubate 2-10 minutes at room temperature and mix several times. Collect beads with a magnetic separator and transfer the supernatant, containing the DNA, into a new tube. If eluate appears brown, repeat collection of the beads. Elution can be improved by repeating these steps or by incubating at 60 °C during elution.
