A 6 month-old male is thought to have sickle cell anemia, and as his pediatrician you want to sequence the gene that may be mutated, causing this disease. You decide to sequence the gene for the beta subunit of hemoglobin. In order to do this you must perform polymerase chain reaction (PCR). You need to know which of the following in order to amplify (PCR) the area of interest of his DNA?
The predicted change in nucleotide sequence
The nucleotide sequence in the areas upstream and downstream to the nucleotides of interest
The predicted change in amino acid sequence of the mutated protein
The change in number of hydrogen bonds within the nucleotide sequence of interest
The predicted change in size of the mutated protein
Correct answer
The nucleotide sequence in the areas upstream and downstream to the nucleotides of interest
Feedback
correct answer: B
Answer: B. The answer is B. Polymerase chain reaction is a method with which to amplify an area of interest of DNA. Normally, specifically designed primers bind upstream and downstream to the area of interest. The sequence in between these primers is amplified and then sequenced to look for specific nucleotide changes that will then predict a possible change in amino acid sequence and protein structure and function. In sickle cell anemia, the beta subunit of hemoglobin is mutated at the 6th codon, and glutamate (wild type) is replaced by valine in the protein. This missense mutation changes protein function and causes sickle cell anemia which is inherited in an autosomal recessive manner.
Choice A is incorrect; the predicted change is unnecessary for performing PCR and sequencing the area of interest. The only known sequences must be where the primers bind to DNA.
Choice C is incorrect; polymerase chain reaction deals with nucleic acids, DNA in this case. Protein sequence is then predicted by changes in the DNA nucleotide sequence.
Choice D is incorrect; predicted changes in numbers of hydrogen bonds may influence the parameters of the PCR assay itself but have no influence on nucleotide or amino acid sequence.
Choice E is incorrect; PCR is done on DNA and thus the predicted size of the protein does not influence the assay itself.