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It does not "self ligate" without ligase, however it can "self ligate" during ligation reaction where ligase is present in the mixture. It is advisable to dephosphorylate digested plasmid vector before ligation to lower the number of colonies with circularized empty plasmid after transformation of the ligation mixture.
#152338 Dephosphorylation
Posted
on 15 March 2013 - 12:59 PM
#152404 Dephosphorylation
Posted
on 16 March 2013 - 12:46 PM
For a single cut, I would dephosphorylate, but you want to use as little of the phosphatase as possible, otherwise you will often find that you will get no clones. CIAP is probably the worst of the phosphatases, though shrimp is pretty bad too, if you use one of the more recent phosphatases from cold organisms, then they tend to be less heat stable and hence more inactivatable.
The vector can not self ligate with ligase, but it is entirely possible that not all of the vector will cut with the RE in the first place.
The vector can not self ligate with ligase, but it is entirely possible that not all of the vector will cut with the RE in the first place.
