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Practice Problems 1. The biochemical pathway below synthesizes two related amino acids D and F. Which components will support the growth of a mutant defective for enzyme 2? Either C or D 2. In a bacterial strain, mutants unable to synthesize an essential compound G were tested with related compounds D, E, and F. The results were as follows, where + means growth. The results are shown in the following table, where + means growth. What is the correct order in which genes act in this pathway? A) 3,4,1,2 B) 1,4,3,2 C) 1,2,3,4 (D) 4,2,1,3 E) 4,3,2,1 3. Two mutants for the rII region of bacteriophage T4 were coinfected and plated E. coli B and K. The table below shows the number of colonies observed in each E. coli strain. Caluclate the map distance in between the two mutations. 0.22 MU 4. A series of rII mutants were crossed and the following frequencies of wild type recombinants were recovered: r1 X r2….0.011 r1 X r3….0.005 r1 X r4….0.003 r2 X r3….0.006 r2 X r4….0.008

r3 X r4….0.002 Indicate the order of the mutants and the map distances between them. Wild type phage that infect E. coli strain K represent only one half of the recombinants. Therefore, double the detected recombination frequencies to estimate the total frequency of recombination. Since one map unit equals 1% recombination, multiply the frequencies by 100. r1 X r2= 2.2; r1 X r3= 1.0; r1 X r4 = 0.6; r2 X r3 = 1.2; r2 X r4=1.6; r3 X r4= 0.4. A map is constructed by assigning mutants a sequence that is additive and compatible with the individual map distances. The two mutants displaying the largest map distance are often at the opposite ends of the map, in this case r1 and r2 r1------(2.2)--------r2. Next find the compatible order of the other mutants: Map is r1----(0.6)------r4----(0.4)------r3-------(1.2)--------r2 5. Pair-wise crosses were made between three mutant strains (numbered 1, 2, and 3). The cultures were plated on strain B and strain K (1/2 on B and 1/2 on K) and the results are given in the table below. Calculate the map distance between each pair of mutations. Distance 1 to 2= 2x10 7 times 2 divided by 2x10 9 = 2x10 -2 = 0.02=2% or 2 map units Distance 1 to 3= 10 8 /10 9 = 10 map units Distance 2 to 3= 1.2x10 8 /10 9 = 12 map units The following table corresponds to the results of pairwise coinfections of E. coli K by T4 bacteriophage mutants for rII the region. + indicates plaques formed. How many different genes are being affected by these mutations? Which deletion mutations and point mutations affect the same genes? Ignore this question, the wrong table was used for this problem 6. Deletion Mutations Point Mutations 1 2 3 4 5 6 7 A 0 + 0 0 + + 0 B + + + 0 + 0 0

C + + + + 0 0 0 D 0 + + 0 + 0 0 E + 0 0 0 + + 0 7. You grow a bacterial culture to a density of 1.2 x10 8 CFU/mL and prepare five 1:10 serial dilutions. You plate 100 µl of the last tube. How many colonies do you expect to see in the plate? Expect to see approximately (1.2 × 10 8 CFU/mL)×(1/10) 5 × 0.1 mL =120 CFU 8. You set up a series of 8 tubes, each containing 9 ml of saline. You add 1 ml of a bacterial suspension to tube 1, mix thoroughly, then transfer 1 ml from tube 1 to tube 2. You repeat this process until you reach tube 8. From each tube, you spread 150 µl onto separate agar plates. After incubation, on the 7 th plate you count 120 colonies. What was the concentration of bacteria in the original culture? 120 colonies x 1/0.15 volume plated = 800 800 x 10 7 (dilution factor) = 8 x 10 9

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