Question

Question 27 6 pts As an undergraduate researcher you have been given the task of identifying what/how many DNA structural forms exist for a particular species of bacterial plasmid. You isolate individual bacterial plasmids from cells and run a gel electrophoresis. You know that there are three possible forms the bacteria DNA can take: superhelical/supercoiled (form1), nicked/open circle/no supercoiling (form II), and linear (form III). a) List the forms from most compact to least compact: b) All three forms have the same number of nucleotides and are, therefore, the same mass. Will these three forms migrate through a gel at the same or different rates? If different rates then which will run the fastest (ie-move the farthest)? Why? HTML Editor BIVAA. IEE 1 x * EE - s o 2 x D : 12pt - Paragraph -

Answer 1

Before jumping to the answer, let's first understand the difference between the three forms.

Form 1 (supercoiled)

Also known as covalently closed-circular (ccc) DNA.

Due to its supercoiled nature, the DNA fragments become smaller in size and highly compact. Therefore, this form experience least frictional resistance from the gel resulting in the faster migration than other conformations.

Form 2 (nicked)

When one of the double strands of the circular supercoiled DNA fragment is broken (due to nick or cut) , the double helix reverts to its normal i.e. uncoiled or relaxed state. This form is called as nicked or open-circular (oc) or relaxed. This conformational change results in the increase in size of the DNA molecule and hence it experiences more frictional resistance from the gel. As a result of this, the migration of open-circular DNA is slower than other conformations.

Form 3 (linear)

When both the strands of the supercoiled DNA fragment is broken, the plasmid becomes linear. As there is no supercoiling, this form is least compact. Because of this conformation, it migrates at normal speed i.e. migrates according to its size.

Based on the above understanding and explanation , the answer can be summarized as below:

Answer a.

The degree of compactness from higher to lower order is

Supercoiled > nicked > linear

Answer b.

The rate of migration of three forms will be (farthest to nearest)

Supercoiled > linear > nicked