Consider the 50 base-pair (bp) DNA sequence below - it is part of a much larger

Question

Consider the 50 base-pair (bp) DNA sequence below - it is part of a much larger DNA molecule: 10 20 3 0 40 50 5 '-CTATGCACTTTGTACAGGGTGCCCGGAGGTTTCTGAACTGTCGACTAGTG-3' 3'-GATACGTGAAACATGTCCCACGGGCCTCCAAAGACTTGACAGCTGATCAC-5' (a) You want to amplify this 5obp region using PCR. You design one 5 nucleotide (nt) primer 5' - CTATG - 3'. What is the sequence of the other 5nt primer needed to amplify this region using PCR? (10 pts) 5 3 (b) There are two alleles of this gene: The "A" allele, which has the sequence shown above The "a" allele, which is the same sequence shown above except that the A/T base pair at position 27 (indicated by a *) has been replaced by a G/C base pair. . . Consider the following four restriction enzyme sites: Mspl 5 '-CCGG-3' Phol 5 '-GGCC-3' Sall 5 '-GTCGAC-3' Smal 3 '-GGCC-5' 3'-CCGG-5 i. Which of the above enzymes would cut DNA from the "A" allele? Circle all that apply. (8 pts)

Explanation / Answer

a. The second primer has the sequence - 5' - CACTA

b. (i) For A allele - MspI, Sal I

    (ii) For a allele - SmaI, MspI, Sal I

c. For 50bp segment - PhoI ( since the specific sequence is absent)

   For 26 and 24 bp - SmaI - a allele

For 50, 26, 24 - a combination of PhoI and MspI

For 42, 8 - Sal I

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