The mutagen 5-bromouracil (5BU) was added to a rapidly dividing culture of wild-type E. coli
cells growing in a liquid medium containing a rich variety of nutrients, including arginine. After
one cell division, the cells were washed free of the mutagen, resuspended in sterile water, and
plated onto master plates containing minimal medium supplemented only with arginine. Plates
were obtained having well-separated colonies, so that each colony derived from just one
progenitor cell. The colonies were then replica-plated from the master plates onto plates
containing minimal medium. One colony that grew in the presence of arginine but failed to grow
on minimal medium was selected from the master plate. The cells of this colony were
suspended in sterile water, and each of 20 tubes containing minimal medium supplemented
with arginine was inoculated with a few cells from this suspension. After the 20 cultures grew to
a density of 108
cells/mL, 0.1 mL from each was plated on plates containing minimal medium.
The following table shows the number of bacterial colonies that grew on each plate.
a. In which stage(s) of this process did mutations occur? What is the evidence that a mutational
event occurred?
b. At each stage where mutations occurred, were the mutations induced or spontaneous? Were
they forward or reverse mutations?
c. At each stage where mutations were recovered, how were they selected for?
d. Though all of the 20 cultures started from a single colony that failed to grow on minimal
medium were treated identically, they produced different numbers of bacterial colonies when
they were plated. Why did this occur?
e. Suppose that 5BU had been added to the medium in the 20 tubes. Would plating the 20
cultures have given the same results? If not, how would they have differed?
f. Supposing that methyl methane sulfonate (MMS) rather than 5BU had been added to the
medium in the 20 tubes, answer the questions given above in part (e).
The mutagen 5-bromouracil (5BU) was added to a rapidly dividing culture of wild-type E. coli
cells growing in a liquid medium containing a rich variety of nutrients, including arginine. After
one cell division, the cells were washed free of the mutagen, resuspended in sterile water, and
plated onto master plates containing minimal medium supplemented only with arginine. Plates
were obtained having well-separated colonies, so that each colony derived from just one
progenitor cell. The colonies were then replica-plated from the master plates onto plates
containing minimal medium. One colony that grew in the presence of arginine but failed to grow
on minimal medium was selected from the master plate. The cells of this colony were
suspended in sterile water, and each of 20 tubes containing minimal medium supplemented
with arginine was inoculated with a few cells from this suspension. After the 20 cultures grew to
a density of 108
cells/mL, 0.1 mL from each was plated on plates containing minimal medium.
The following table shows the number of bacterial colonies that grew on each plate.
a. In which stage(s) of this process did mutations occur? What is the evidence that a mutational
event occurred?
b. At each stage where mutations occurred, were the mutations induced or spontaneous? Were
they forward or reverse mutations?
c. At each stage where mutations were recovered, how were they selected for?
d. Though all of the 20 cultures started from a single colony that failed to grow on minimal
medium were treated identically, they produced different numbers of bacterial colonies when
they were plated. Why did this occur?
e. Suppose that 5BU had been added to the medium in the 20 tubes. Would plating the 20
cultures have given the same results? If not, how would they have differed?
f. Supposing that methyl methane sulfonate (MMS) rather than 5BU had been added to the
medium in the 20 tubes, answer the questions given above in part (e).
