Chapter 12, Problem 9P

Two wild-type fragments of human genomic DNA from the long arms of two nonhomologous acrocentric chromosomes are shown in the accompanying diagram. On both fragments, the centromere-to-telomere direction is left-to-right. The red lines numbered 1, 2, and 3 indicate the positions of breakpoints that could be caused by ionizing radiation. Certain combinations of these breaks could produce chromosomal rearrangements when the broken pieces are stitched back together by DNA repair systems.

You will want to diagnose the presence of the rearrangements by using PCR. In every case that follows, one of the primers you will use for PCR is primer A, whose sequence is:

5′ TCGATTCCGGAAAGCT 3′

a.	Which two of the breakpoints could yield a deletion?
b.	Write the sequence and polarity of a 16-base primer that, in conjunction with primer A, will allow you to diagnose the presence of a deletion in a patient’s genomic DNA. The evidence should be positive, not negative. (That is, you will see a PCR product whose presence and/or size is specific to the deletion; the absence of a band is not informative.)
c.	Which two of the breakpoints could yield an inversion?
d.	Write the sequence and polarity of a 16-base primer that, in conjunction with primer A, will allow you to diagnose the presence of an inversion in a patient’s genomic DNA. Your answer should show the primer that would yield the longest possible diagnostic PCR product. Your primer cannot cross any of the red lines.
e.	Which two of the breakpoints could yield a reciprocal translocation? (Two possible answers exist; you need only write one.)
f.	Write the sequence and polarity of a 16-base primer that, in conjunction with primer A, will allow you to diagnose by PCR the presence of a reciprocal translocation in a patient’s genomic DNA. This reciprocal translocation can be stably inherited through meiosis without its loss or any chromosomal breakage. Again, the evidence should be positive, not negative.
g.	Is the translocation you described in parts (e) and (f) Robertsonian? Answer yes or no, and explain.