Denaturation and Renaturation of DNA:
In this article we will discuss about the Denaturation and Renaturation of DNA.
Denaturation of the DNA:-
The process of formation of single stranded DNA from a double stranded helical DNA by heating or denaturating agents is called a denaturation of DNA.
Denaturation refered to the separation of 2strands of a DNA.
Denaturation can be done by the following denaturating agents:-
a) Denaturation by Temperature
b) Denaturation by Chemical agents
c) Effects of pH denaturation
(a) Denaturation by Temperature:-
If a DNA solutions is heated to approximately 90°c Or above there will be enough kinetic energy to denature the DNA completely causing it to separate in to single strand. This denaturation is accelerated by chemical reagents like urea and formaldehyde.
The chemical enhance the aqueous solubility of the purine and pyrimidine group. This separation of double helix is called melting as it occurs abrupt at a certain characteristics temperature called denaturation. Temperature or melting temperature (TM).
If several sample of DNA are melted, it is found that the TM is highest for those DNA that contain the highest proportion of G=C.
The relationship between Tm and G= C contain arises due to formation of three hydrogen bond between guanine and cytosine when base paired where as adenine and thymine form only 2hydrogen bonds.
b) Denaturation by chemical agents:-
Denaturation of DNA double helix can be brought about by certain chemical agents such as urea & formaldehyde. These chemical reagents enhance the aqueous solubility of the purine and pyrimidine of urea DNA can be completed denaturation by 95% formaldehyde at room temperature only.
c) Effect of pH denaturation:-
Denaturation also occurs at acidic and alkaline solution in which ionic change of purine and pyrimidine bases can occure.
At acidic solution the pH value 2 to 3 the amino group binds with proton and DNA double helix is disrupted.
Renaturation of DNA:-
The process of formation of double stranded DNA from a single stranded DNA upon pulling is called renaturation of DNA.
When preparation of double stranded DNA are denatured and allowed to renature, the rate of renaturation can give variable information. About the complexity of the DNA. It there are repetitive. Sequence the DNA it shows less complexity in comparison to it’s totall length of all sequence are unique. A sample with a highly repetitive sequence will renature rapidly, while complex sequence will renature slowly.
If 1killobase (1kb) DNA fragments are denatured by heating above it’s Tm and then renatured at a temperature 10°c below the Tm and monitored either by decrease in absorbance at 260nm or by passing sample at the interval through a column of hydroxylapatite, which retains only double standard DNA and estimating how much of the sample retain.