we transformed plasmids in two two conditions, one with ligase and one without ligase.
both of them showed colonies.
The plate treated with ligase is bound to ligate as the plasmid is circular and the cells will grow even in an antibiotic marker medium.
One person says the colonies are due to incomplete digestion of the plasmid and some of the undigested plasmids get transformed as a circular DNA and colonies appear.
though the plasmids which are digested cannot religate as there is no ligase for the phospho- diesterase activity and even if it gets transformed in the bacterial cells, the DNA will be chopped off by exonuclease activity.
the other person says the plasmid will re-ligate inside the bacterial cell after transformation due to the presence of internal DNA ligases and hence colonies appear.
what i feel is plasmid should get re-ligated due to the conformational change (with sticky ends) and/or internal ligases and/or incomplete digestion. I feel there should be one more control experiment done with alkaline phosphatase to assess the DNA ligase activity.
So please clear my doubt that can the plasmid get re-ligated itself without adding DNA ligase? if so, how?
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Edited by Arun, 04 June 2009 - 02:32 AM.