Double Digestion Experiment

1) Present the gel electrophoresis image of plasmid digestion experiment with the correct label. Describe the results of the experiment and list down possible problem that you observe based on the electrophoresis image. Plasmid P1-1 and P1-2 undergo single digestion. Plasmid P1-3 undergoes double digestion.

Single digestion is only one restriction enzyme which has been used to digest a DNA.

Double digestion is there are 2 restriction enzymes which have been used to digest a DNA.

Based on the result, in the gel electrophoresis, P1-1 has form two bands as it has 2 restriction sites.

For P1-2, there is only one band formed as there is only one restriction site.

For P1-3, there are three bands formed as there are 3 restriction sites which have been cut by restriction enzymes.

Based on the gel electrophoresis, the higher the concentration of plasmid DNA, the brighter the colour of the bands. The size of P1 is smaller than P2. The distance of the bands in P1 is

This include, the time taken for incubation of the enzymes is typically 1 hour. The unit definition of restriction enzymes is based on a 1-hour incubation period. Each enzyme has different incubation period. For example, enzyme such as ECoRI need maximum period. 2hrs. If more than 2 hours, it will lead to the star activity. Enzyme should be the last component which added to reaction before incubate it under 37°C. This is because the activity of the enzymes can be maintained at optimal temperature, 37°C. The mix components through pipetting the reaction mixture should be invert up and down slowly so that the mixture components are equalised. It can also follow by quick spinning in a micro centrifuge to ensure the components in the microtube are mixed equally. Enzymes should be keep in the ice when it is not store in the freezer because heat from the surrounding can cause denaturation of the enzymes and eventually cause them