Experiment 3 - Chris Stanton Leah Thursday Rm 41 Experiment...

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Chris Stanton Leah, Thursday Rm 41 Experiment 3: Characterizations of Nonsense Mutations in E.Coli Purpose : The purpose of this experiment was to mate a lac - Hfr strain of E.coli with different F - strains that had a deletion of the lac-pro region and specific amber suppressor genes in order to determine if the lac - mutants contained missense or nonsense mutations and whether the nonsense mutations could be suppressed by known amber suppressors. For an amber suppressor to work, the genetic mutation causing lac - had to be due to a nonsense mutation with the codon UAG, and the inserted amino acid had to restore function to the protein. Controls: The controls in this experiment were extremely important. The obvious controls, which were the individual strains diluted with water, allowed us to be sure that the mated strains had the characteristics required for the experiment. If any plate had growth, it meant that the strain was not what it should have been and that the colonies on the mated plates would not necessarily be the exconjugates. For example, the HfrC lac - strain was plated on min + strep and min + nal. Although the lac - strain would have been able to grow on min plates which contain glucose as the carbon source, the lac - Hfr could not grow on the min + strep and min + nal because it was sensitive to both of these substances. Similarly, the CSH54 could not grow on min + strep because, although it would have had the lactose it needed and it was resistant to Streptomycin, it was pro - and the min + strep plate did not provide the proline necessary for growth. The CSH55 and CSH56 strains contained the same
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