PNK Treatment of DNA Ends protocol

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(New page: <html> <h2>Solutions/reagents:</h2><ul type="circle"><li> <a name="PNK stock">PNK stock <i><br><tab><div style="margin-right: 600px;">(10, 000 U/ml)</div></i></a></li><li>T4 ligase buffer<...)
Current revision (03:15, 20 November 2009) (view source)
 
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<h2>Solutions/reagents:</h2><ul type="circle"><li> <a name="PNK stock">PNK stock <i><br><tab><div style="margin-right: 600px;">(10, 000 U/ml)</div></i></a></li><li>T4 ligase buffer</li><li>substrate</li></ul><h2>Equipment:</h2><ul type="circle"><li>Incubator</li><li>Reaction tubes</li></ul><h2>Steps:</h2><ol><p><li><b><font size=3>Reaction Mix (10 µl)</font></b><br>Use the following table as a checklist for preparing the reaction:<br><br><table border cellpadding=5 rules=all frame=void bordercolor=#357EC7><thead><tr><td>&nbsp;</td><td><font color=#357EC7>PNK stock</font></td><td><font color=#357EC7>T4 ligase buffer</font></td><td><font color=#357EC7>substrate</font></td></tr></thead><tbody><tr><td><font color=#357EC7>Reaction Mix</font></td><td><b><b><font color=#357EC7>1 µl</font></b></td><td><b><b><font color=#357EC7>1 µl</font></b></td><td><b><b><font color=#357EC7>8 µl</font></b></td></tr></body></table></li></p><p><li><b><font size=3>Reaction Conditions</font></b><br>Incubate at <b><font color=#357EC7>37°C</font></b> for <b><font color=#357EC7>30 mins</font></b>.<br>Incubate at <b><font color=#357EC7>65°C</font></b> for <b><font color=#357EC7>20 mins</font></b>.<br>Store at <b><font color=#357EC7>4°C</font></b>.<br></li></p></ol>
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<h2>Solutions/reagents:</h2><ul type="circle"><li> <a name="PNK stock">PNK stock <i><br><tab><div style="margin-right: 600px;">(10, 000 U/ml)</div></i></a></li><li>T4 ligase buffer</li><li>substrate</li></ul><h2>Equipment:</h2><ul type="circle"><li>Incubator</li><li>Reaction tubes</li></ul><h2>Steps:</h2><ol><p><li><b><font size=3>Reaction Mix (10 µl)</font></b><br>Use the following table as a checklist for preparing the reaction in reaction tube (1):<br><br><table border cellpadding=5 rules=all frame=void bordercolor=#357EC7><thead><tr><td>&nbsp;</td><td><font color=#357EC7>PNK stock</font></td><td><font color=#357EC7>T4 ligase buffer</font></td><td><font color=#357EC7>substrate</font></td></tr></thead><tbody><tr><td><font color=#357EC7>Reaction Mix</font></td><td><b><b><font color=#357EC7>1 µl</font></b></td><td><b><b><font color=#357EC7>1 µl</font></b></td><td><b><b><font color=#357EC7>8 µl</font></b></td></tr></body></table></li></p><p><li><b><font size=3>Reaction Conditions</font></b><br>Incubate at <b><font color=#357EC7>37°C</font></b> for <b><font color=#357EC7>30 mins</font></b>.<br>Incubate at <b><font color=#357EC7>65°C</font></b> for <b><font color=#357EC7>20 mins</font></b>.<br>Store the tube <b><font color=#357EC7>on ice</font></b>.<br></li></p></ol><p><b>TOTAL TIME REQUIRED FOR THE COMPLETION OF THE PROTOCOL :<font color=#357EC7>~ 50 mins</font></b></p>
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Current revision

Solutions/reagents:

Equipment:

  • Incubator
  • Reaction tubes

Steps:

  1. Reaction Mix (10 µl)
    Use the following table as a checklist for preparing the reaction in reaction tube (1):

     PNK stockT4 ligase buffersubstrate
    Reaction Mix1 µl1 µl8 µl

  2. Reaction Conditions
    Incubate at 37°C for 30 mins.
    Incubate at 65°C for 20 mins.
    Store the tube on ice.

TOTAL TIME REQUIRED FOR THE COMPLETION OF THE PROTOCOL :~ 50 mins

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