Has anyone ever experienced that colonies disappear from plates after transformation? I cloned a small fragment using NEB cloning kit and transformed into XL1blue. I got few colonies, kept the plate in fridge for two days and surprised by seeing that colonies disappeared ! What could be the reasons?
Dear khandaker
Sounds like the plasmid somehow conferred transient antibiotic resistance, and then the either the plasmid has disappeared or the resistance has gone. you will have to repeat the transformation anyway. I would recommend that you not add more DNA than 5% of the cell volume - too much can inhibit the transformation process.
best wishes.
This is a very strange outcome indeed. Above unfortunately copied and pasted their answer from another site that had a similar question asked a few years ago (http://www.protocol-online.org/biology-forums-2/posts/27937.html). While I'm not big into naming and shaming this isn't really what the research gate questions section is for.
I would suggest reading through the above conversation as it may have some helpful tips.
I would suggest conducting a positive control. Transform an un-edited plasmid into XL1 blue and see if the colonies disappear. This would help identify if it's a transformation inefficiency or if the recombinant plasmid you made is unstable.
Agreed with Colman answer.
Better transform an unedited. This will give you an insight if you are missing something during your work.
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