Question

Case Study 12-2 A 7-month-old child suffered from sleep apnea and difficulty breathing. Respiratory infections and allergies were ruled out. From the family history, the physician suspected congenital congestive hypoventilation syndrome (CCHS). CCHS results, in part, from abnormal function of the PHOX2B gene product caused by a triplet-repeat expansion. The expansion of the triplet GCN in the DNA sequence (N is any base) results in the insertion of additional alanines in the Phox2B protein. The symptoms of the syndrome include interruptions in breathing during sleep and other abnormalities in the autonomic nervous system. A blood sample from the patient was tested for the GCN expansion by PCR using primers that flanked the location of the potential expansion. The amplicons were resolved by agarose gel electrophoresis. <Insert Fig 12-26> gcagcagcagcggeggcggecgeggcagcggcogeggcggcageggcagcggcogcagct PHOX2b exon 3 A 1 2 3 4 5 6 7 8 9 10 11 12 PAGE 23 20 -(Normal) D 400- 300. (Normal) 200. Agarose (rapid test)

1. The PCR procedure can be performed with fluorescently labeled primers and polyacrylamide gel electrophoresis (PAGE). What would be the advantage of this procedure over agarose gel analysis?
   1. PAGE allows for higher resolution than the agarose gel procedure.
   2. The PAGE procedure is faster than that using agarose gel electrophoresis.
   3. The labeled primers are less expensive because they are used at a lower concentration in the reaction mix.
   4. The PAGE procedure is less prone to contamination

Answer 1

Ans: DNA fragments smaller than 100 bp are more effectively separated using polyacrylamide gel electrophoresis. Unlike agarose gels, the polyacrylamide gel matrix is formed through a free radical driven chemical reaction. These thinner gels are of higher concentration, are run vertically and have better resolution.

Hence option A is the advantage of PAGE over agarose gel electrophoresis.