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- A prototrophic strain (his" arg" lac') was used as a donor to transform an auxotrophic strain (his arg lac'). Initial transformants are isolated on minimal medium + histidine + arginine + lactose - glucose. i. What genotypes will grow on this medium? ii. These colonies are replicated to minimal medium + histidine, and 50% of the original colonies grow. What genotypes will grow on this medium? iii. The original colonies are also replicated to minimal medium + arginine, and 10% of the colonies grow. What genotypes will grow on this medium? iv. The original colonies are also replicated to minimal medium. No colonies grow. Based on this information, what genotypes will grow on minimal medium + histidine and on minimal medium + arginine? What is the relative gene order for his, arg, and lac? Which two genes are closer? Explain your answer.In Escherichia coli, four different Hfr strains, derived from the same F* strains, were mated with F strains auxotrophic for a number of nutritional requirements (Arg Bio Cys Trp Gal His Lac Mal Xyl Leu Met). Matings were interrupted at various intervals and cells were plated on minimal medium supplemented with particular nutrients to test for gene transfer. The following results show the time of entry for each of the genes in four different Hfr strains. Table 1: Time-of-Entry Mapping Data* Hfr strains Genes lac" his" arg' bio 9. Cys gal" trp' 5 11.5 2.5 mal" xyl" leu met Hfr 1 Hfr 2 Hfr 3 Hfr 4 6.5 3.5 11 15 15 4 6. 17.5 5 14.5 3 20 * The numbers denote the number of minutes elapsed before a gene enters the F cells. Draw a circular map of E. coli chromosome.Given what we've discussed in class, what will be most likely outcome if you conjugate an streptomycin resistant ampicillin sensitive methionine auxotroph E. coli strain (engineered to be pir+) that is F- with a streptomycin sensitive non-HFR methionine prototroph strain that is F- and RP4+ but contains pUC18? Colonies on minimal media + ampicillin +streptomycin plates No colonies on minimal media +ampicillin +streptomycin plates
- In a generalized-transduction experiment, phages arecollected from an E. coli donor strain of genotype cys+leu+ thr+ and used to transduce a recipient of genotypecys- leu- thr-. Initially, the treated recipient populationis plated on a minimal medium supplemented with leucine and threonine. Many colonies are obtained.a. What are the possible genotypes of these colonies?b. These colonies are then replica plated onto threedifferent media: (1) minimal plus threonine only, (2)minimal plus leucine only, and (3) minimal. Whatgenotypes could, in theory, grow on these three media?c. Of the original colonies, 56 percent are observed togrow on medium 1, 5 percent on medium 2, and nocolonies on medium 3. What are the actual genotypes ofthe colonies on media 1, 2, and 3?d. Draw a map showing the order of the three genes andwhich of the two outer genes is closer to the middle geneA microbial geneticist isolates a new mutation in E. coliand wishes to map its chromosomal location. She usesinterrupted-mating experiments with Hfr strains andgeneralized-transduction experiments with phage P1.Explain why each technique, by itself, is insufficient foraccurate mapping.By conducting conjugation experiments between Hfr and recipientstrains, Wollman and Jacob mapped the order of many bacterialgenes. Throughout the course of their studies, they identified severaldifferent Hfr strains in which the F-factor DNA had been integratedat different places along the bacterial chromosome. A sample of theirexperimental results is shown in the following table:Explain how these results are consistent with the idea that thebacterial chromosome is circular?
- Three pairs of bacterial cells with the given genotypes undergo conjugation. Place match the genotype of each cell after conjugation to its initial genotype. F+ x F Hfr F- F' F- Answer Bank F F F+ Hfr What is the role of the F-factor in conjugation? It contains genes necessary for replication of the donor's F plasmid. can occur. It allows auxotrophic bacterial cells to survive on minimal medium so that conjugation It contains genes that force recombination between the donor and recipient chromosomes. It contains genes necessary for the formation of the pilus. O It degrades the chromosome of the recipient cell after conjugation.a. Bacteriophage P22 was used in generalized transduction experiments to infect the Salmonella typhimurium donor strains described in the table below. The resulting phage lysates were then used to infect the S. typhimurium recipient strains listed in the table. In each cross, a phenotype was selected for one of the three genetic markers studied (str, aceA, thrA), and then replicates were performed to select the corresponding recombinants for the other two markers. The results are given in the following table: Recipient strain Selected phenotype Selected recombinants Donor strain str thrA aceAt thrA str aceA+ strs thrA+ aceA thrA+ str aceA Str Ace+ Str ThrA ThrA+ ThrA ThrA+ Ace Ace+ str: gene involved in streptomycin resistance, aceA: gene involved in the use of acetate as a carbon source, thrA: gene involved in the biosynthesis of threonine. Number 60 40 95 5 10 90 What are the selective media used in these three transduction experiments, on the one hand to obtain the selected…A pure culture of an unknown bacterium was streaked onto plates of a variety of media. You notice that the colony morphologyis strikingly different on plates of minimal media with glucose compared to that seen on trypticase soy agar plates. How can you explain these differences in colony morphology? Also, describe what happens when a nonsense mutation is introduced into the gene encoding transposase within a transposon and why is it more likely that insertions or deletions will be more detrimental to a cell than point mutations?
- Austin Taylor and Edward Adelberg isolated some new strains of Hfr cells that they then used to map several 200 mal+ genes in Escherichia coli by using interrupted conjugation. 150 In one experiment, the researchers mixed cells of Hfr strain AB-312, which were xyl* mtl* mal* met* and sensitive to phage T6, with F strain AB-531, which was xyl mtl mal met and resistant to phage T6. The 100 mt/+ cells were allowed to undergo conjugation. At regular intervals, the researchers removed a sample of cells and 50 met+ interrupted conjugation by killing the Hfr cells with phage T6. The F cells, which were resistant to phage T6, survived and were then tested for the presence of 0. 20 40 60 80 100 genes transferred from the Hfr strain. The results of this experiment are shown in the graph. Time of sampling (minutes) On the basis of these data, give the order of the xyl, mtl, mal, and met genes on the bacterial chromosome and the minimum distances between them in minutes. The origin of transfer is…Bacteriophage P22 was used in generalised transduction experiments to infect the Salmonella typhimurium donor strains described in the table below. The resulting phage lysates were then used to infect the recipient strains of S. typhimurium recipient strains listed in the table. In each cross, a phenotype was selected for one of the selected for one of the three genetic markers studied (str, aceA, thrA), and were made to select the recombinants corresponding to the other two markers. markers. The results are given in the following table: Strain I donor str thrA aceA thrA str aceA+ Strain recipient strs thrA+ aceA thrA str aceA Phenotype selected Str Ace+ Str recombinants selected ThrA ThrA ThrA ThrA Ace Ace Number 60 40 95 5 10 90 str: gene involved in streptomycin resistance, aceA: gene involved in the use of acetate as a carbon source, thrA: gene involved in threonine biosynthesis. 1) What are the selective media used in these three transduction experiments? to obtain the selected…Bacteriophage P22 was used in generalized transduction experiments to infect the Salmonella typhimurium donor strains described in the table below. The resulting phage lysates were then used to infect the S. typhimurium recipient strains listed in the table. In each cross, a phenotype was selected for one of the three genetic markers studied (str, aceA, thrA), and then replicates were performed to select the corresponding recombinants for the other two markers. The results are given in the following table: Recipient strain Selected phenotype Selected recombinants Donor strain str thrA aceA+ thrA str aceA+ strs thrA+ aceA thrA+ str aceA Str Ace+ Str ThrA ThrA+ ThrA ThrA+ Ace Ace str: gene involved in streptomycin resistance, aceA gene involved in the use of acetate as a carbon source, thrA: gene involved in the biosynthesis of threonine. Number 60 40 95 5 10 90 Determine the order of the genes and draw a genetic map showing this order