Difference between revisions of "Troubleshooting/Ligation"

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<b>No colonies</b>
 
<b>No colonies</b>
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<p>
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If you know your cells are working well, there are a few common ligation problems that might be happening with your reaction.
  
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<ol><li><b>Uncut plasmid:</b> It's possible that your destination backbone was not cut well during your digest. </li>
+
<ol><li><b>Uncut insert:</b> It's possible that your insert was not cut well during your digest. </li>
 
<li><b>Ligase didn't work:</b>
 
<li><b>Ligase didn't work:</b>
  

Revision as of 19:40, 7 May 2015

Ligation Troubleshooting

Now that you know you transformation efficiency and it's above 1 x 108 CFU/µg DNA, we can work on other possible problems if you're still not getting great results from your cloning. This page is focused on common problems students have with ligations.

No colonies

If you know your cells are working well, there are a few common ligation problems that might be happening with your reaction.

  1. Uncut insert: It's possible that your insert was not cut well during your digest.
  2. Ligase didn't work: