Question
Selecting the recombinants becomes cumbersome when antibiotic is inactive- explain.###How can we distinguish between recombinant and non recombinant plasmid? Illustrate.
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Answer
→ Selection of recombinants due to inactivation of antibiotics is a cumbersome procedure because it requires simultaneous plating on two plates having different antibiotics.
→ Therefore, alternative selectable markers have been developed which differentiate recombinants from non-recombinants on the basis of their ability to produce colour in the presence of a chromogenic substrate.
→ In this, a recombinant DNA is inserted within the coding sequence of an enzyme, ẞ-galactosidase. This results into inactivation of the gene for synthesis of this enzyme, which is referred to as insertional inactivation.
→ The presence of a chromogenic substrate gives blue coloured colonies if the plasmid in the bacteria does not have an insert.
→ Presence of insert results into insertional inactivation of the ẞ-galactosidase gene and the colonies do not produce any colour, these are identified as recombinant colonies.
→ Therefore, alternative selectable markers have been developed which differentiate recombinants from non-recombinants on the basis of their ability to produce colour in the presence of a chromogenic substrate.
→ In this, a recombinant DNA is inserted within the coding sequence of an enzyme, ẞ-galactosidase. This results into inactivation of the gene for synthesis of this enzyme, which is referred to as insertional inactivation.
→ The presence of a chromogenic substrate gives blue coloured colonies if the plasmid in the bacteria does not have an insert.
→ Presence of insert results into insertional inactivation of the ẞ-galactosidase gene and the colonies do not produce any colour, these are identified as recombinant colonies.
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