BICD100 2008 midterm 2 KEY for 2010

BICD100 2008 midterm 2 KEY for 2010 - 2008 BICD100 Midterm...

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1 2008 BICD100 Midterm 2 KEY (mean 107/150 points) 1.) In the course of studying heavy metal metabolism in the bacterium Escherichia plumbum , you identify a mutant ( alc - ) that can convert lead into gold (wild type E. plumbum can’t do this). Before you can patent your strain, you need to characterize the gene that causes this remarkable alchemy phenotype. The alc - mutant strain you isolated is F - and carries 6 other mutations, his - , lys - , trp - , phe - , met - and neo r . Thus, this strain requires addition of histidine, lysine, tryptophan, phenylalanine and methionine to the medium in order to grow, and is also resistant to the antibiotic neomycin. A map showing the locations of these marker mutations is shown to the right (assume this chromsome has total of 100 minutes on it, like the E. coli chromosome). To map the alc gene you take advantage of three different Hfr strains that are wild type for all genes (all 3 are his + , lys + , trp + , phe + , met + , neo s and alc + ). You mix your F - mutant and Hfr wild type strains, disrupt the mating mixtures at different times up to 40 minutes, and plate the mixtures on media containing neomycin, histidine, lysine, tryptophan, phenylalanine and methionine. You score the resulting colonies for his, lys, trp, phe, met and alc phenotypes. a. What is the purpose of selecting for neomycin resistance (one sentence)? (5 pts) This kills the Hfr donor cells (which are neomycin sensitive), allowing determination of which Hfr donor alleles were transferred to F - recipients (which are neomycin resistant). You observe the intial appearance of Hfr donor alleles in F - recipients at the following times (- means that transfer was not observed at any of the timepoints tested): transferred from>> Hfr#1 Hfr#2 Hfr#3 his + - - 30 min lys + 30 min 10 min - trp + 7 min 33 min - phe + 19 min 21 min - met + - - 11 min alc + 8 min 34 min - b. Show on the chromosome map above the location of the F insertion in each Hfr strain (show its location in minutes and use an arrowhead to indicate the direction of transfer). (9 pts) See above c. Also show on the chromsome map above the approximate location of the alc gene. (6 pts) See above (alc is within about a minute of trp but we can’t tell from this data which side of trp it is on) Problem continues on next page…
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2 To map alc more precisely, you cross wild type Hfr strain 1 with the F - mutant strain and thirty mintues later plate the disrupted mating mixture on media containing neomycin, histidine, lysine, tryptophan, and methionine but not phenylalanine. 500 of the resulting neo r phe + colonies are then tested for their ability to grow in the absence of tryptophan and for their ability to turn lead into gold, with the following results: grew without tryptophan and couldn’t turn lead to gold: 173 grew without tryptophan but could turn lead into gold: 2 could not grow without tryptophan and couldn’t turn lead into lead into gold: 29 could not grow without tryptophan but could turn lead into gold: 296 d. Based on this information, draw a map of the region where the alc gene maps - not the whole circular
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