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Question
The rate of migration of DNA in an agarose gel is affected by which of the following factor(s)?
I. Concentration of DNA in the sample loaded on gel
II. Conformation of DNA molecule
III. Size of the DNA fragment
IV. Source from where DNA has been isolated
The rate of migration of DNA in an agarose gel is affected by which of the following factor(s)?
I. Concentration of DNA in the sample loaded on gel
II. Conformation of DNA molecule
III. Size of the DNA fragment
IV. Source from where DNA has been isolated
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Solution
The correct option is B II and III
Gel electrophoresis is a laboratory technique used for the separation of a mixture of DNA (a negatively charged molecule) fragments of varying sizes on agarose gel (a natural biopolymer) matrix under the influence of electric field.
Agarose gel is a polymer of agarobiose composed of alternating units of D-galactose and 3,6-anhydro-L-galactopyranose. The cross linking of the monomers creates pores in the gel whose size is determined by the degree of cross linkage. Wells are dug on the gel into which the sample DNA is pipetted out.
On application of electric field start, the DNA molecules start migrating towards the oppositely charged terminal (anode). However, the speed of migration is affected by the size and conformation of the DNA molecules.
- Size: Size of a double stranded DNA fragment is determined by the number of base pairs present in it. Keeping all other experimental parameters constant, the linear DNA molecules of smaller size or containing lesser number of base pairs can percolate swiftly through the pores present in the gel compared to the one which is larger in size containing greater number of base pairs. This makes the smaller sized DNA molecules migrate faster or cover a longer distance towards the anode over stipulated time.
- Conformation: Conformation refers to the folding of the linear chains of molecules in the three dimensional space. Keeping all other experimental parameters constant, a DNA molecule with a compact conformation would migrate easily through the pores compared to a molecule with an extended conformation. This will affect the rate of migration. For example, a supercoiled DNA is more compactly packed than a linear one, hence, migrates with a higher speed.
On the other hand, the concentration of DNA in the sample loaded per well does not have any effect on the rate of migration. However, it is a determinant of the thickness of the band during visualisation. A lesser quantity of the sample would allow intercalation of a lesser number of ethidium bromide molecules. This would result in a comparatively less prominent band.
Source of the DNA sample might require different DNA isolation protocols. However, it does not affect the rate of migration of DNA molecules on the agarose gel during electrophoresis.
Gel electrophoresis is a laboratory technique used for the separation of a mixture of DNA (a negatively charged molecule) fragments of varying sizes on agarose gel (a natural biopolymer) matrix under the influence of electric field.
Agarose gel is a polymer of agarobiose composed of alternating units of D-galactose and 3,6-anhydro-L-galactopyranose. The cross linking of the monomers creates pores in the gel whose size is determined by the degree of cross linkage. Wells are dug on the gel into which the sample DNA is pipetted out.
On application of electric field start, the DNA molecules start migrating towards the oppositely charged terminal (anode). However, the speed of migration is affected by the size and conformation of the DNA molecules.
- Size: Size of a double stranded DNA fragment is determined by the number of base pairs present in it. Keeping all other experimental parameters constant, the linear DNA molecules of smaller size or containing lesser number of base pairs can percolate swiftly through the pores present in the gel compared to the one which is larger in size containing greater number of base pairs. This makes the smaller sized DNA molecules migrate faster or cover a longer distance towards the anode over stipulated time.
- Conformation: Conformation refers to the folding of the linear chains of molecules in the three dimensional space. Keeping all other experimental parameters constant, a DNA molecule with a compact conformation would migrate easily through the pores compared to a molecule with an extended conformation. This will affect the rate of migration. For example, a supercoiled DNA is more compactly packed than a linear one, hence, migrates with a higher speed.
Source of the DNA sample might require different DNA isolation protocols. However, it does not affect the rate of migration of DNA molecules on the agarose gel during electrophoresis.
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