Plated Replica

Plated Replica confused with this question? help?????? pBR322 (which has tetracycline and ampicillin resistance genes) is digested with the restriction endonuclease Pst1, annealed with foreign DN...

Plated Replica

confused with this question?

help??????

pBR322 (which has tetracycline and ampicillin resistance genes) is digested with the restriction endonuclease Pst1, annealed with foreign DNA, ligated with DNA ligase and transformed into Escherichia coli. The Pst1 site resides within the tetracycline resistance gene. The transformed clones are isolated and replica plated on tetracycline and ampicillin agars. Which of the following patterns of antibiotic resistance will indicate that the clones carry:

[1] a re-circularised plasmid? ……………………………………………………………
and

[2] a recombinant plasmid incorporating the foreign DNA? .................................

(i) The clone is resistant to both tetracycline and ampicillin
(ii) The clone is resistant to tetracycline and sensitive to penicillin
(iii) The clone is sensitive to tetracycline and resistant to ampicillin
(iv) The clone is sensitive to both tetracycline and penicillin

There are 2 separate answers

1 recircularized plasmid: just means the cut plasmid PstI site re-annealed and you have amp & tet resistant pBR322 that you started with, So colonies that grow on amp and tet have this plasmid.
(i) The clone is resistant to both tetracycline and ampicillin

2 recombinant plasmid: the tet(R) gene is disrupted by the foreign DNA inserted in the PstI site, so plasmid still confers resistance to amp, but not tet. If you pick colonies that grow on amp but not tet you should recover the recombinant plasmid.
(iii) The clone is sensitive to tetracycline and resistant to ampicillin