A plasmid that encodes resistance to ampicillin and tetracycline is digested wit

Question

A plasmid that encodes resistance to ampicillin and tetracycline is digested with the restriction enzyme PstI which cuts the plasmid at a single site in the ampicillin-resistance gene. The DNA is then aneealed with PstI digest of human DNA, ligated and used to transform E.Coli cells. a) what antibiotic-resistance phenotypes will be found on the plate  b) what antibiotic woiuld you put in an agar plate to ensure that the cells of a bacterial colony contain the plasmid withou an insert?    c) which phenotype will indicate the presence of plasmids that contain human DNA fragments?

Explanation / Answer

Bacteria have existed on Earth for three billion years or so and have become adept at protecting themselves against toxic chemicals. Antibiotics have been in clinical use for a little more than 6 decades. That antibiotic resistance is now a major clinical problem all over the world attests to the success and speed of bacterial adaptation. Mechanisms of antibiotic resistance in bacteria are varied and include target protection, target substitution, antibiotic detoxification and block of intracellular antibiotic accumulation. Acquisition of genes needed to elaborate the various mechanisms is greatly aided by a variety of promiscuous gene transfer systems, such as bacterial conjugative plasmids, transposable elements and integron systems, that move genes from one DNA system to another and from one bacterial cell to another, not necessarily one related to the gene donor. Bacterial plasmids serve as the scaffold on which are assembled arrays of antibiotic resistance genes, by transposition (transposable elements and ISCR mediated transposition) and site-specific recombination mechanisms (integron gene cassettes).The evidence suggests that antibiotic resistance genes in human bacterial pathogens originate from a multitude of bacterial sources, indicating that the genomes of all bacteria can be considered as a single global gene pool into which most, if not all, bacteria can dip for genes necessary for survival. In terms of antibiotic resistance, plasmids serve a central role, as the vehicles for resistance gene capture and their subsequent dissemination. These various aspects of bacterial resistance to antibiotics will be explored in this presentation.

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