Question

3. (7) Suppose that you have selected E. coli that can grow only when the medium is supplemented with cysteine. You isolate one mutant that you believe is defective in a new gene require for cysteine biosynthesis - cysX .
You have two strains of bacteria:
Hfr cysX - gal+ his+ lac+ pur+ trp+ ton s
F - cysX+ gal - his - lac - pur - trp - ton R
A) What supplements would you need to add to minimal media to recover all cysX+, lac+, tonR recombinants? Would you add glucose?
B) What recombinant genotypes would you recover on plates containing only minimal media, glucose, hisitidine, purines, and T1 phage?
C) When you conduct an interrupted mating experiment with the Hfr and F - strains, Ton R
colonies with the following phenotypes first appear at the times shown in the table below:

0 Phenotype Minutes cys+ cys- 25 gal+ 17 his+ 38.5 lac+ 9.5 pur+ 14.5 trp+ 25

Draw a bacterial chromosome and indicate the relative position of each of the markers and the cysX gene in the Hfr strain. Be sure to label distances between markers.

D) Based on the results in (C), you decide to also perform mapping of the cysX gene using P1 cotransduction. P1 phage grown on an E. coli strain with the genotype cysX - ton R trp+ are used to infect a recipient strain of E. coli with the genotype cysX+ ton S trp - . These bacteria are then plated on complete media in the presence of T1 phage. Two hundred ton R transductants are scored for their genotype with respect to the other markers. Here are the results:

What is the recombination frequency between trp and cysX ?
E) What is the recombination frequency between ton and cysX ?
F) What is the order of the three genes? Indicate the distance between them.

Answer 1

A) The supplements supplied should be Galactose/Glucose, Histidine, Purine, Tryptophan and T1 phage. Yes I would add glucose.

B) The obtained genotype will be Gal- His- Pur- tonR CysX+ Lac+ Trp + .

C)

2.5 et

D) Recombinant Frequency between Trp and Cys X = 10/200 = 5%

Order of the genes is CysX tonR Trp.