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Question
CBSEENBI12002022
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Explain the role(s) of the following in biotechnology.
(a) Restriction endonuclease
(b) Gel-electrophoresis
(c) Selectable markers in pBR322

Solution

(a) Restriction Endonucleases These are the bacterial enzymes that cut dsDNA into fragments after recognising and binding to the specific nucleotide sequences, known as recognition site. These enzymes are used to form recombinant molecules of DNA, composed of DNA from different sources.

(b) Gel-Electrophoresis is the technique which allows the visualisation of separated fragments of DNA on an agarose gel matrix.
Since, the DNA fragments are negatively charged molecules, they separate and move towards the anode (+ ve) under the influence of an electric field. DNA fragments are separated on the basis of their size through the sieving effect provided by the gel.
(c) Selectable Markers in pBR322, an E. coli cloning vector has two antibiotic resistance genes, i.e. for ampicillin and tetracycline. Antibiotic resistance is used as selectable marker, i.e. it helps in identification and selection of transformants. For example, when a foreign DNA is ligated at the site of tetracycline resistance (tet r ,) gene in pBR322, the recombinant plasmid will lose tetracycline resistance due to insertional inactivation of foreign DNA, but can still be selected out from non-recombinants by placing the transformants on ampicillin containing medium. The transformants growing on ampicillin containing medium, are then transferred on tetracycline containing medium. The recombinants will grow on ampicillin containing medium but not on tetracycline one whereas non-recombinants grow on both.

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