Chapter 14, Problem 32P

Scientists who study amino acid biosynthesis pathways want to isolate auxotrophic bacteria. A technique called penicillin enrichment makes this task easier. This procedure starts by exposing a liquid culture of wild-type (prototrophic) bacteria growing in rich (complete) medium to a chemical mutagen. After this treatment, the cells are centrifuged to remove the liquid and the mutagen. The pellet of cells at the bottom of the centrifuge tube is now resuspended in medium that lacks one amino acid (in this example, cysteine) but contains penicillin. Subsequently, the bacteria are poured onto a filter that concentrates them and allows them to be washed free of the penicillin. The living bacteria retained on the filter are highly enriched for cysteine auxotrophs.

a.	Given what you know about the action of penicillin, explain why this enrichment occurs.
b.	Penicillin enrichment is not a selection, because the drug does not kill 100% of the prototrophs. The cells on the filter thus need to be screened for cysteine auxotrophy. How would the scientists perform this screen?
c.	If the starting strain contained a pen gene on a plasmid, would this scheme still enrich for auxotrophs? Explain.