question_answer 54)

    For selection of recombinants, insertional inactivation of antibiotic marker has been super coded by insertional inactivation of a marker gene coding for a chromogenic substrate. Give reasons.

Answer:

                  In selection of recombinants due to inactivation of antibiotics, the transformed cells are first plated on the antibiotic plate which has not been insertionally inactivated (i.e., ampicillin)and incubated overnight for growth of transformants. For selection of recombinants, these transformants are replica-plated on second antibiotic(say, tetracycline) plate (which got inactivated due to insertion of gene), Non-recombinants grow on both the plates (one carrying ampicillin and the other carrying tetracycline) while recombinants will grow only on ampicillin plate. This entire exercise is labourious and takes more time (two overnight incubation) as well However, if we choose insertional inactivation of a marker that produces colour in the presence of a chromogenic compound, we can distinguish between the recombinants and non-recombinants on a single medium plate (containing one antibiotic and the chromogenic, compound) after overnight growth.