Difference between revisions of "Team:Paris Bettencourt/Protocols/gel-purification"

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       <li>Add 3 volume of QG buffer (100mg of gel ~ 100ul) in the tube and incubate for 10 min at 50°C (or 10 min on a vortex).
 
       <li>Add 3 volume of QG buffer (100mg of gel ~ 100ul) in the tube and incubate for 10 min at 50°C (or 10 min on a vortex).
 
       <li>Add one volume of isopropanol and mix.
 
       <li>Add one volume of isopropanol and mix.
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       <li>vortex the tube for 2 min at 14 000 rpm.
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       <li>Centrifugate the tube for 2 min at 14 000 rpm.
 
       <li>Carefully pipette the supernanant in a QIAquick column.
 
       <li>Carefully pipette the supernanant in a QIAquick column.
 
       <li>Put the column in a 2ml microcentrifuge tube and centrifuge it for 1 min at 14 000 rpm.
 
       <li>Put the column in a 2ml microcentrifuge tube and centrifuge it for 1 min at 14 000 rpm.
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       <li>Throw the supernatant.
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       <li>Discard the flow-through.
 
       <li>Add 750 uL of PE buffer in the column and centrifuge for 1 min and discard flow-through.
 
       <li>Add 750 uL of PE buffer in the column and centrifuge for 1 min and discard flow-through.
 
       <li>Centrifuge again for 2 min and put the column in a new 1.5uL microcentrifuge tube.
 
       <li>Centrifuge again for 2 min and put the column in a new 1.5uL microcentrifuge tube.
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Latest revision as of 14:50, 29 August 2015

Gel-Purification protocol with the QIAquick® Gel Extraction Kit
  • Excise the DNA band from the gel as precisely as possible and put it in a micro-centrifuge tube.
  • Add 3 volume of QG buffer (100mg of gel ~ 100ul) in the tube and incubate for 10 min at 50°C (or 10 min on a vortex).
  • Add one volume of isopropanol and mix.
  • Centrifugate the tube for 2 min at 14 000 rpm.
  • Carefully pipette the supernanant in a QIAquick column.
  • Put the column in a 2ml microcentrifuge tube and centrifuge it for 1 min at 14 000 rpm.
  • Discard the flow-through.
  • Add 750 uL of PE buffer in the column and centrifuge for 1 min and discard flow-through.
  • Centrifuge again for 2 min and put the column in a new 1.5uL microcentrifuge tube.
  • Elute the DNA by putting 50uL of water in the middle of the column and let it stand for 2 min.
  • Centrifugate for 2min at 10 000 rpm