During agarose gel electrophoresis of DNA, shorter DNA molecules typically migra

Question

During agarose gel electrophoresis of DNA, shorter DNA molecules typically migrate faster than longer molecules. When closed-circular DNA molecules of the same size possess different linking numbers, these DNA molecules will also separate during agarose gel electrophoresis. A sample of purified plasmid DNA of uniform length was separated on an agarose gel and stained, as shown below. In addition to a strongly stained band (B), the gel shows a faintly stained band (A) that migrated a much shorter distance. Select the statement that explains why two bands appeared in the migration pattern. O Band B DNA is more supercoiled than band A DNA. o Topoisomerase I supercoiled band B DNA and topoisomerase II supercoiled band A DNA. Relaxed DNA is more susceptible to DNA damage, resulting in two populations of DNA size groups. Band B DNA is less supercoiled than band A DNA. Band B DNA is negatively supercoiled, whereas band A DNA is O positively supercoiled. Incorrect The two bands shown in the migration pattern do not result from damage to the DNA sample. How does supercoiling affect DNA migration through an agarose gel? How would the addition of DNA ligase and DNA gyrase to the plasmid sample prior to electrophoresis alter the migration pattern? The migration pattern will change from discrete bands to a smear of many species.

Explanation / Answer

Hi
When plasmids are run on the gel, we often get 3 bands. One belongs to supercoiled DNA which is also called cccDNA, runs faster on the gel. The another band which lags behind is the ocDNA or open circle DNA, this experience higher friction and moves slowly on the gel. Thus the B DNA in the example, is more supercoiled than A DNA.

The supercoiling occurs when there is a nick in one of the DNA strands. The addition of DNA ligase, can acutally join these broken strands in the supercoiled DNA (band B), and DNA gyrase can introduce more negative supercoils. Thus the band A will disappear.

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