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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The plasmid (pBR322) has resistance genes for both ampicillin and tetracycline. Show more…
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A plasmid that encodes resistance to ampicillin and tetracycline is digested with the restriction enzyme BamHI, which cuts the plasmid at a single site within the tetracycline resistance gene. The DNA is then annealed with a BamHI digest of human DNA, ligated with DNA ligase, and used to transform E. coli cells. a) What antibiotic would you include in an agar plate to ensure that the cells of a bacterial colony that grow on the plate contain the plasmid? b) What two antibiotic-resistance phenotypes (with respect to ampicillin and tetracycline resistance) will be found on the plate? c) Which phenotype (from part b) will indicate the presence of plasmids that contain the desired human DNA fragments?
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