An experiment was performed on isolation, cloning and transformation of plasmid DNA. The objective of this experiment was to successfully clone a kanamycin resistance gene into the multiple cloning site of a pUC18 plasmid and then to transform competent cells with the plasmids.

Although both lanes in the agarose gel, contain plasmid DNA, why does the DNA not appear to be in the same location in both lanes ? How would you verify that the transformed cells actually contain the amp/kan plasmid that was used for transformation ?

see explanation.

Step-by-step explanation

Although both lanes in the agarose gel, contain plasmid DNA, but the DNA does not appear to be in the same location in both lanes because in gel electrophoresis the DN fragments get separated according to there size. The large DNA fragments wil move short distance in gel and the short or circular DNA will move comparatively long distance.

after digesting with an specific restriction enzyme which cut site present in the MSC of pUC18 will produce only one large fragment that cant be able to move in gel so no band will appear but in case of pkan plasmid no fragment will produce hence that transformed circular plasmid will move some distance through gel and will show only one band.