Team:Grenoble/Biology/Protocols/DNA extract

From 2012.igem.org

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                 <tr><td><div class="petit">The Membrane Binding Solution is considered as harmful that is why in
                 <tr><td><div class="petit">The Membrane Binding Solution is considered as harmful that is why in
                 case of liquid on the skin or in the eyes, it must be washed with soap
                 case of liquid on the skin or in the eyes, it must be washed with soap
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                 <tr><td><div class="petit">The centrifuge must be balanced before being switched on. In case of
                 <tr><td><div class="petit">The centrifuge must be balanced before being switched on. In case of
                 dysfunction of the device, it must not be used.</div></td></tr>
                 dysfunction of the device, it must not be used.</div></td></tr>
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                 <tr><td><div class="petit">The use of membrane wash solution is made up with ethanol. When using the centrifuge,
                 <tr><td><div class="petit">The use of membrane wash solution is made up with ethanol. When using the centrifuge,
                 the lid is closed. Then it can happen an accumulation of ethanol gas. It is important to open the lid once
                 the lid is closed. Then it can happen an accumulation of ethanol gas. It is important to open the lid once

Revision as of 07:25, 3 September 2012

iGEM Grenoble 2012

Project

DNA extraction from Agarose gel

Goal

Extract DNA from Agarose gel.

Protocol

Following are the instructions from the NucleoSpin® Extract II Kits User Manual.

  1. Excise DNA fragment with a scalpel.

  2. Add 200 µL Buffer NT for each 100 mg agarose gel. Incubate sample at 50°C until the gel slices are dissolved.

  3. SAFETY AND USEFUL RECOMMANDATION
    The Membrane Binding Solution is considered as harmful that is why in case of liquid on the skin or in the eyes, it must be washed with soap and water. However the quantity is small therefore the risk is minimum. The gel is also full of ethidium bromide. This product is not volatile, and its melting point is around 240°C, therefore there is almost no risk of inhalation.

  4. Place a NucleoSpin® Extract II column into a 2 mL collecting tube and load the sample. Centrifuge for 1 min at 11,000 x g.

  5. SAFETY AND USEFUL RECOMMANDATION
    The centrifuge must be balanced before being switched on. In case of dysfunction of the device, it must not be used.

  6. Discard flow-through and place the column back into the collecting tube.

  7. Add 600-700 µL buffer NT3. Centrifuge for 1 min at 11,000 x g. Discard flow-through and place the column back into the collecting tube.

  8. SAFETY AND USEFUL RECOMMANDATION
    The use of membrane wash solution is made up with ethanol. When using the centrifuge, the lid is closed. Then it can happen an accumulation of ethanol gas. It is important to open the lid once manipulations are finished to evacuate this product.

  9. Centrifuge for 2 min at 11,000 x g. You can also incubate the column for 2-5 min at 70°C prior to elution.

  10. Place the column into a new clean 1.5 mL eppendorf tube. Add 15-50 µL elution buffer NE, incubate at room temperature for 1 min. Centrifuge for 1 min at 11,000 x g.