A certain E. coli gene, gene X, codes for a protein that has histidine (His) at amino acid position
100. The complete protein is 300 amino acids long. E. coli genomic DNA is amplified in a PCR
reaction using primers that bind near opposite ends of the gene. The PCR product is 950 bp long.
The PCR product is digested with the restriction endonuclease BadI and the fragments are
separated on an agarose gel. The BadI DNA fragments are 300, 400, and 250 bp long.
In a mutant E. coli strain, a single base pair substitution has caused a missense mutation such
that His100 is replaced by arginine (Arg) in the protein. When DNA from this strain is amplified, a
950 bp product is again obtained, but when that is digested with Badl only two fragments are
obtained, 700 and 250 bp.
BadI recognizes and cleaves the DNA sequence 5′ G T G C A C3′
a. What amino acid is at position 99 of the protein in the original strain?
b. What amino acid is at position 99 of the protein in the mutant strain?
c. Can you determine the identity of the original His100 codon? If so, which is it?
d. Can you determine the identity of the Arg100 codon? If so, which is it?
e. Was the mutation a transition or a transversion?
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