E X E R C I S E S
I. Interpreting Your Gel
1. Observe the photograph of the stained gel containing your sample and those from other students. Orient the photograph with the sample wells at the top. Interpret the band(s) in each lane of the gel:
a. Scan across the photograph to get an impression of what you see in each lane. You should notice that virtually all student lanes contain one or two prominent bands.
b. Now locate the lane containing the pBR322/BstN I markers. Working from the well, locate the bands corresponding to each restriction fragment: 1,857 bp, 1,058 bp, 929 bp, 383 bp, and 121 bp (may be faint).
c. The amplification product of 440 bp should roughly align with the 383 bp marker.
d. It is common to see a second band lower on the gel. This diffuse (fuzzy) band is "primer dimer," an artifact of the PCR reaction that results from the primers overlapping one another and amplifying themselves. Primer dimer is approximately 50 bp, and should be in a position ahead of the 121 bp marker.
e. Additional faint bands, at other positions on the gel, occur when the primers bind to chromosomal loci other than mt control region and give rise to "nonspecific" amplification products.
2. How would you interpret a lane in which you observe primer dimer, but no 440-bp band?
3. The mt control region mutates at approximately 10 times the rate of nuclear DNA. Propose a biological reason for the high mutation rate of mt DNA.
4. The high mutability of the mt genome means that it evolves more quickly than the nuclear genome. This makes the mt control region a laboratory for the study of DNA evolution. However, can you think of any drawbacks to this high mutation rate?
5. There are numerous insertions of mt DNA into nuclear chromosomes. Notably, scientists recently discovered a 540-bp fragment of the mt control region that inserted into chromosome 11 approximately 350,000 years ago. Would you expect any difference in the mutation rates of the control region sequence in the mt genome versus the chromosome 11 insertion? What implication does this have in the study of human evolution?
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