7.) (18 Points).
Based upon proflavin-induced mutants in the rII locus of phage T4 and pseudorevertants
(intragenic suppressors) of these original mutants, Crick et al deduced that the genetic
code must be a multiple of three nucleotides.
Wild-type T4 phage produce smooth plaques (W) on E. coli B, but rII mutants form
rough plaques (R) on E. coli B. rII mutants cannot form plaques on E. coli K while wildtype T4 form W plaques. A partial sequence of the rII region of T4 is shown below.
(Only the coding strand is shown for simplicity). Mutants (designated as FC followed by
a number) are shown below the sequence. Some mutants, like FC40 and FC 32 insert a
base while others such as FC18 and FC88 delete a base.
5’ A T T C A A G T G G T A A T T A C A C C C A A A 3’
FC40
FC18
FC32
FC88
FC33
+A
-A
+A
-C
+C
The phenotypes of phage with single mutations or combinations of mutations are shown
in the following table. “R” means the phage makes rough plaques. “W” indicates that
the phage makes wild-type plaques.
A). In the last column in the table on the next page, explain the reason why each
combination produces R or W plaques. Include an explanation that explains the
phenotype at the level of mRNA translation. (A genetic code dictionary is included at the
end of the exam). (12 Points)
B). Brenner isolated a mutant of E. coli K12 that allowed the FC18 and FC32
combination to make wild type plaques. What is the most likely explanation for this
result? (3 Points).
[The mutant probably contained an ochre suppressor that reads UAA codons].
C). What does this work suggest about the amino acid sequence of the above region of
the rII gene? (3 Points).
[The amino acids in this region of the protein are not important for function of the
rII gene.]
Mutation(s)
FC40
Plaques on
E. coli B
R
FC88
R
FC18
R
FC32
R
FC40 + FC88
W
Reason for Phenotype
FC40 and FC88 are a (+) and (-)
combination and the phenotype becomes
wild-type indicating that the reading frame
between the sites does not contain a
nonsense codon.
FC40 + FC32
R
Both FC40 and FC32 are (+) additions.
The prediction is that the translational
reading frame is still wrong.
FC18 + FC32
R
FC18 and FC32 are (-) and (+) but the (-)
creates an in frame TAA codon before the
(+) addition. Thus there is translation
termination and the mutant phenotype.
FC40 + FC32 + FC33
W
The three (+) additions restore the correct
reading frame. If you draw out the
sequence you will see that there are no
termination codons in the new mRNA
made by the triple mutant.