Assume that rfp and its promoter pBAD were cloned from the pKAN-R plasmid into the pARA plasmid in a sample of host bacteria. How would you verify which bacteria possessed this new hybrid plasmid (hint: think of the effect that arabinose has on the pARA plasmid)? Remember that both this hybrid plasmid and the pKAN-R plasmid both have rfp, so you'll need to distinguish the two as well (hint: what happens when bacteria are plated on media containing antibiotics?).
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Step 1: Plate the host bacteria on media containing both kanamycin and ampicillin antibiotics. Show more…
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A plasmid (pBR322) that encodes resistance to ampicillin and tetracycline is digested with the restriction enzyme BamHI, which cuts the plasmid at a single site in the tetracycline resistance gene. The DNA is then annealed with BamHI digest of human DNA, ligated, and used to transform E. coli cells. You would put ampicillin in the agar plate to ensure that the cells of the bacterial colony contain the plasmid. How can you select the clones that contain the human DNA? Explain what you would do and what the results would be if the clone contained human DNA.
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