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Possible to lose a pGEM insert ? - (Apr/23/2009 )


i have a quick question, can an insert (about 500 bp) "fall out" of pGEM once ligated into it ?

am i stupid for thinking it can be lost !




You could do an agarose gel electrophoresis, if you see a band from 3500bp you have the right one, if you see only a band from 3000, you lose it.

Or you could you cut the plasmid with EcoRI, for verify the integrity of your construction.


It can happen, depending on the type of insert. Some sequences like IRES or similar sequences can be lost.