Which of the following is/are NOT the step(s) used in separation and isolation of DNA fragments by gel electrophoresis technique:
(A) Staining with ethidium bromide
(B) Exposure to UV Radiation
(C) Cutting DNA fragments into small pieces by using restriction endonuclease
(D) Denaturation of DNA fragments
Choose the correct answer from the options given below:
Answer & explanation
Correct answer: option 1
The correct answer is Option (1) → (D) only
(A) Staining with ethidium bromide — A step. Makes the DNA bands visible.
(B) Exposure to UV Radiation — A step. Stained bands show up under UV light.
(C) Cutting DNA fragments into small pieces by using restriction endonuclease — A step. Cutting by restriction enzymes gives the fragments that are separated.
(D) Denaturation of DNA fragments — Not a step. DNA is not denatured for separation on agarose gel.
NCERT Extract:
Restriction Enzymes – “… Separation and isolation of DNA fragments : The cutting of DNA by restriction endonucleases results in the fragments of DNA. These fragments can be separated by a technique known as gel electrophoresis. Since DNA fragments are negatively charged molecules they can be separated by forcing them to move towards the anode under an electric field through a medium/matrix. Nowadays the most commonly used matrix is agarose which is a natural polymer extracted from sea weeds. The DNA fragments separate (resolve) according to their size through sieving effect provided by the agarose gel. Hence, the smaller the fragment size, the farther it moves. … The separated DNA fragments can be visualised only after staining the DNA with a compound known as ethidium bromide followed by exposure to UV radiation (you cannot see pure DNA fragments in the visible light and without staining). You can see bright orange coloured bands of DNA in a ethidium bromide stained gel exposed to UV light (Figure 9.3). The separated bands of DNA are cut out from the agarose gel and extracted from the gel piece. This step is known as elution.”
