Name: UCLA ID#

Section:

INTRODUCTION TO GENETICS LS4 Summer 2010 08/27/2010 MIDTERM II

Please circle your TAs name: ANSHU MOHSIN RON

Instructions: 1. This exam has 7 pages. Before starting, make sure you have all the pages. 2. Clearly write your name on every page. Be sure to write clearly. Only legible answers will be graded. 3. Do not write on the backs of pages. We will not grade them. 4. All books, notes, and cell phones (turned off) must be put in zipped backpacks. 5. Calculators may be used. NO GRAPHING CALCULATORS ARE ALLOWED!

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NAME: 1. (16 Points) Benzers Experiments A. Mutations rII1 and rII2 are known to fall into the rIIA and rIIB cistrons (genes) respectively. Mutations rII1 and rII2 are used to infect E. coli B at high m.o.i and the progeny phage collected. These phage are then used to infect strain K at very low multiplicity of infection (m.o.i.). The resulting phage are diluted and plated on both E. coli B and E. coli K. Do you expect more plaques on strain B, on Strain K, an equal amount on both or is it impossible to tell? (circle one) E. coli B E. coli K Equal on both Can't tell

B. As in (Part A), cells of E. coli B are infected simultaneously with phage strains rII1 and rII2 at a high moi. The progeny phage are collected and used to infect cells of E. coli K. This infection is also carried out at a high m.o.i., and the resulting phage are collected. These progeny are diluted and plated on both E. coli B and E. coli K. Do you expect more plaques on strain B, on strain K, an equal amount on both or is it impossible to tell? (circle one) E. coli B E. coli K Equal on both Can't tell

C. E. coli strain B is doubly infected with two rII mutants of phage T4. 0.1 ml of a 107 dilution of the progeny is plated on E. coli B and 0.1 ml of a 103 dilution of the progeny is plated on E. coli K. 25 plaques appeared on strain B, 7 on strain K. Calculate the recombination frequency between these two mutations. # Recombinants/Total Progeny = 2 x K/B 2 (7 x 103)/ 2.5 x 108 = 5.6 x 10-5 = 5.6 x 10-5 x 100 = 0.0056% Note: If they simply put 5.6 x 10-5 then that fine because that is the recombination frequency. Highlighted above is the Percent Recombination (full credit also).

NAME: 2. (16 Points) Conjugation: Hfr x FAn Hfr strain of genotype: phe+ arg+ eryr strs is mated with a recipient strain of genotype: phe- arg- erys strr. Exconjugants are plated on a medium containing Str (streptomycin), Ery (erythromycin), Arg (arginine), and Phe (phenylalanine). The following results are obtained for 400 colonies isolated from these plates and tested for growth on various media: a. MM + Ery: b. MM + Ery + Arg: c. MM + Ery + Phe: 293 302 318

a) Write a list of possible genotypes, and indicate the number of individuals in each genotype. eryr arg+ phe+ = 293 eryr arg+ phe- = 318 293 = 25 eryr arg- phe+ = 302 293 = 9 eryr arg- phe- = 400 (293 + 25 + 9) = 73

b) What is the gene order? Include the position of the origin of transfer. Among these three markers, ery is the last one to be transferred. eryr-------arg-----pheO

c) Calculate the recombination frequencies. arg-phe = (25 + 9)/400 x 100 = 8.5 mu ery-arg = (73+9)/400 x 100 = 20.5 mu ery-phe = 8.5 + 20.5 = 29 mu

NAME: 3. (6 Points) Selecting for the most exconjugants Consider the cross: Hfr str-s arg- met+ gal+lys+ x F- str-r arg+ met- gal- lys- where the genetic map is shown below (the < indicates the Origin of transfer in the Hfr strain).

gal arg

met

str lys
Circle the media below which would select for the most exconjugants. a) Minimal media b) Minimal media + streptomycin c) Minimal media + methionine d) Minimal media+ arginine e) Minimal media + galactose + streptomycin f) Minimal media+ streptomycin+ methionine g) Minimal media + methionine + galactose + lysine h) Minimal media + galactose + arginine + lysine + methionine i) Minimal media + streptomycin + galactose + methionine + lysine 4. (10 points) Complementation/Recombination. Choose the correct answer in each case: (circle one) (A) If one wants to know if two different rII point mutations lie at exactly the same site (nucleotide pair), one should: i. coinfect E. coli K with both mutants. If phage are not produced they lie at the same site. ii. coinfect E. coli K with both mutants. If phage are produced they lie at the same site. iii. coinfect E. coli B with both mutants and plate the progeny phage on both E. coli B and E. coli K. If plaques appear on B but not K they lie at the same site. iiii. coinfect E. coli B with both mutants and plate the progeny phage on both E. coli B and E. coli K. If plaques appear on K but not B they lie at the same site. (B) If one wants to know if two different rII point mutations lie in the same cistron, one should: i. coinfect E. coli K with both mutants. If phage are not produced they lie in the same cistron. ii. coinfect E. coli K with both mutants. If phage are produced they lie in the same cistron. iii. coinfect E. coli B with both mutants and plate the progeny phage on both E. coli B and E. coli K. If plaques appear on B but not K they lie in the same cistron. iiii. coinfect E. coli B with both mutants and plate the progeny phage on both E. coli B and E. coli K. If plaques appear on K but not B they lie in the same cistron. 4

NAME: 5. (18 points) Transduction (Show your work) Using P22 as a generalized transducing phage grown on an arg+ met+ pro+ bacterial donor, a recipient strain of genotype arg- met- pro- is infected and incubated. Afterwards, transductants for arg+ and met+ are selected individually in experiments I and II respectively. Experiment I arg+ met+ pro+ 0% arg+ met+ pro- 25% arg+ met- pro+ 27% arg+ met- pro- 48% Experiment II arg+ met+ pro+ 0% arg+ met+ pro- 25% arg- met+ pro+ 0% arg- met+ pro- 75%

A. What media was used for the arg+ selection in Experiment I. (circle the correct answer) 1. minimal media 2. minimal media + arg 3. minimal media + met 4. minimal media + pro 5. minimal media + arg + met 6. minimal media + arg+ pro 7. minimal media + met + pro 8. minimal media + arg + met+ pro 9. complete media 10.none of the above B. Calculate co-transduction frequencies and between the three genes. arg and met have a co-transduction frequency of 25% arg and pro have a co-transduction frequency of 27% met and pro have a co-transduction frequency of 0% C. The correct gene order is: (circle the correct answer) arg-met-pro met-arg-pro arg-pro-met

D. Draw the recombination event required to give the arg+ met+ pro- progeny in Experiment #1.

NAME: 6. (12 points) Neurospora You have isolated 100 auxotrophic strains in neurospora. None of these 100 strains can grow on minimal media, but all 100 can grow on minimal media plus methionine and arginine. Further analysis shows that 30 can grow on minimal media plus arginine and 20 can grow on minimal media plus methionine. Fill in the number of strains that have the following phenotype. met- argmet+ argmet- arg+ met+ arg+ total 50 30 20 0 100

7. 10 points (Show Your Work) The h mutant of phage T4 grow in both E. colis and E. colir strains whereas the wild type (h+) phage T4 can grow only on the E. colis strain. You want to determine the recombination frequency between two h mutations (called h1 and h2). The E. colis strain is coinfected (at high m.o.i.) with a wild type phage stock and with a phage that contains both mutant alleles of h. h1- h2- x h1+ h2+

When the progeny is titered it is determined that 5 x 108 pfu/ml of the progeny phage stock can grow on E. colis but only 3 x 108 pfu/ml can grow on E. colir. What is the recombination frequency between the two h mutations. 5 x 108 = total # of phages = # wt non-recombinants + # h1-h2- non-recombinants + + # h1- recombinants + # h2- recombinants 3 x 108 = (# h1-h2- non-recombinants + # h1- recombinants + # h2- recombinants) Therefore, # wt non-recombinants = 5 x 108 - 3 x 108 = 2 x 108 and # of h1-h2- non-recombinants is also 2 x 108 The recombination frequency= 108/5 x 108 = 1/5 = 20%

NAME:

8. (12 Points) Biochemical Genetics: One gene One Enzyme It is known that compounds A, B, C, D are biochemical intermediates in the synthesis of compound E. You perform a mutagenesis for auxotrophic mutants that require compound E to be supplied in the medium for growth and identify five classes of mutants, 1-5. Identify the order of the intermediate compounds in the biochemical pathway (A-E) and identify the step in the pathway affected by each class of mutant (1-5).

A Mutant 1 2 3 4 5 3 Precursor C + + + + 4

B + + + -

C + 1 D

D + + -

Tryptophan + + + + + 2 5 A Tryptophan