Team:Oxford/protocols/QIAquick Gel Extraction

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<li>After growing cell cutlers overnight in LB, resuspend pelleted cells in 250 μl Buffer P1 and transfer it into a microcentrifuge tube.<BR><BR>
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<li>Add 250 μl of Buffer P2 and mix by inverting the tube 6 times. The cell suspension should turn blue.<BR><BR>
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<li>Add 350 μl of Buffer N3 and mix the suspension immediately by inverting 6 times.<BR><BR>
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<li>Centrifuge for 10 minutes at 13000 rpm (or 17900 x g). Apply the supernatants to a spin colums<BR><BR>
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<li>Centrifuge again for 30-60 seconds and discard the flow-through.<BR><BR>
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<li> Wash the spin column by adding 500 μl of the PB buffer and centrifuge for 30-60s.<BR><BR>
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<li>Discard the flow-through and wash the spin column again by adding 750 uL of PE buffer. Centrifuge for 1 minute.<BR><BR>
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<li>Discard the flow-through and centrifuge again for 1 minute. Now place the column in a 1.5ml microcentrifuge tube.<BR><BR>
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<li>Add 50 μl of the EB buffer to the centre of the spin column. Let stand for 2 minutes and centrifuge for 1 minute.<BR><BR>
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Revision as of 13:33, 23 July 2014

QIAquick Gel Extraction

↩ Back to other protocols.

  1. After growing cell cutlers overnight in LB, resuspend pelleted cells in 250 μl Buffer P1 and transfer it into a microcentrifuge tube.

  2. Add 250 μl of Buffer P2 and mix by inverting the tube 6 times. The cell suspension should turn blue.

  3. Add 350 μl of Buffer N3 and mix the suspension immediately by inverting 6 times.

  4. Centrifuge for 10 minutes at 13000 rpm (or 17900 x g). Apply the supernatants to a spin colums

  5. Centrifuge again for 30-60 seconds and discard the flow-through.

  6. Wash the spin column by adding 500 μl of the PB buffer and centrifuge for 30-60s.

  7. Discard the flow-through and wash the spin column again by adding 750 uL of PE buffer. Centrifuge for 1 minute.

  8. Discard the flow-through and centrifuge again for 1 minute. Now place the column in a 1.5ml microcentrifuge tube.

  9. Add 50 μl of the EB buffer to the centre of the spin column. Let stand for 2 minutes and centrifuge for 1 minute.

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