Read the following paragraph and answer the given questions. Repeated amplification is achieved by the use of a thermostable DNA polymerase (isolated from a bacterium, Thermus aquaticus), which remain active during the high temperature induced denaturation of double stranded DNA. 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. The separated bands of DNA are cut out from the agarose gel and extracted from the gel piece. The convention for naming these enzymes is the first letter of the name comes from the genus and the second two letters come from the species of the prokaryotic cell from which they were isolated, e.g., EcoRI comes from Escherichia coli RY 13. In EcoRI, the letter 'R' is derived from the name of strain. Roman numbers following the names indicate the order in which the enzymes were isolated from that strain of bacteria. Stanley Cohen and Herbert Boyer accomplished this in 1972 by isolating the antibiotic resistance gene by cutting out a piece of DNA from a plasmid which was responsible for conferring antibiotic resistance. Since the DNA is enclosed within the membranes, we have to break the cell open to release DNA, RNA, protein, polysaccharides etc. The RNA can be removed by treatment with ribonuclease whereas proteins can be removed by treatment with protease. Other molecules can be removed by appropriate treatments and purified DNA ultimately precipitates out after the addition of chilled ethanol. |
In agarose gel electrophoresis, the separated DNA fragments can be visualized with the help of compound known as: |
Ethidium chloride Rubidium bromide Ethidium bromide Cesium bromide |
Ethidium bromide |
The correct answer is Option (3) -Ethidium bromide The process of cutting DNA using restriction endonucleases results in the formation of DNA fragments. These fragments can be separated using a technique called gel electrophoresis. Since DNA fragments are negatively charged molecules, they can be separated by applying an electric field, causing them to move towards the anode through a medium or matrix. Agarose, a natural polymer extracted from seaweeds, is commonly used as the matrix. The DNA fragments separate based on their size due to the sieving effect provided by the agarose gel. Smaller fragments move farther compared to larger fragments. The separated DNA fragments can be visualized only after staining the DNA with a compound called ethidium bromide, followed by exposure to UV radiation. Without staining, the pure DNA fragments cannot be seen in visible light. After staining, you can observe bright orange-colored bands of DNA in an ethidium bromide stained gel exposed to UV light. Once the DNA fragments are separated, they can be cut out from the agarose gel and extracted from the gel piece. This step is known as elution. The purified DNA fragments obtained through elution are then used in constructing recombinant DNA by joining them with cloning vectors. |