For selection of recombinants, insertional inactivation of antibiotic marker has been superseded by insertional inactivation of a marker gene coding for a chromogenic substrate. Give reasons.
For selection of recombinants, insertional inactivation of antibiotic marker has been superseded by insertional inactivation of a marker gene coding for a chromogenic substrate. Give reasons.
Selection of recombinants due to inactivation of antibiotics is a laborious process as it requires:
(i) a vector with two antibiotic resistance marker
(ii) preparation of two kinds of media plate, with one antibiotic each.
Transformed cells are first plated on that antibiotic plate which has not been insertional inactivated (ampicillin) and incubated overnight for growth of transformants. For selection of recombinants, these transformants are Replica plated on second antibiotic (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 laborious and takes more time (two overnight incubation) as well. However, if we choose the second option (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.
Hence I would choose a marker which produces a coloured compound but gets inactivated due to insertion of foreign DNA.
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