Does DNA Replication Follow the Conservative, Semiconservative, or Dispersive Model?
Introduction—The Article and Phenomenon Under Study
When Watson and Crick published their double-helix model of DNA structure in 1953, they noted that the
complementary pairing of nitrogenous bases suggested, in theory, a possible mechanism for replication.
However, in practice, it was difficult to test their idea. In innovative and careful research with E. coli bacteria,
investigators Meselson and Stahl used a novel way to label DNA and were able to confirm the mechanism of
DNA replication. This research, from the following paper, is the subject of this week’s discussion:
M. Meselson and F. W. Stahl, The replication of DNA in Escherichia coli, Proceedings of the National
Academy of Sciences USA 44:671–682 (1958).
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC528642/pdf/pnas00686-0041.pdf
A. About the Article
1. State the last names of the authors of the article and the name of the university at which they worked.
2. What is the name of the scientific journal in which the article was published? In what year was it
published?
3. This research was aided by grants from what organizations? (See the notes at the end of the article, page
682, before the references.)
4. Specialized vocabulary: Write a brief definition of each term.
density-gradient centrifugation
generation time
isotope
B. About the Study
5. The investigators “anticipated that a label which imparts to the DNA molecule an increased density”
(Introduction, paragraph 1) might help reveal how atoms in parental DNA are distributed to progeny
DNA molecules. What label did the investigators use to be able to detect small density differences
among molecules?
1
6. When bacteria cells are lysed (broken open) and the contents are placed in a cesium chloride mixture
and spun in a centrifuge, the contents of the solution will be sorted by density. Figure 2a presents the
results of separation of DNA composed of N14 from DNA composed of N15 by density-gradient
centrifugation. The left side of the photograph is the part of the centrifuge tube closest to the axis of
rotation (top of the tube), and the right side is where material has spun farthest from the center of
rotation by centrifugal force (bottom of the tube). Which would have greater density, DNA composed of
N14 or DNA composed of N15? Therefore, in a photograph such as in Figure 2a, which is the location of
the higher-density material in each sample—to the left or to the right?
7. DNA absorbs ultraviolet light, so UV light photography can be used to make DNA visible. Figure 1
presents 13 images, each a photograph of ultraviolet absorption, indicating the presence of DNA at
certain locations in the sample in a centrifuge tube. At time 0, before centrifugation, the dark band
covers nearly the entire length of the sample in the tube, from left to right. This means that before
centrifugation, the DNA molecules are scattered throughout the mixture of cell contents in the centrifuge
tube and have not yet been sorted according to density. (a) After 10.7 hours of centrifugation, the dark
band is limited to about what fraction of the length of the centrifuge tube? (b) After what amount of time
(hours) in the centrifuge did the band begin to narrow? (c) After 36.5 hours of centrifugation, the dark
band is located about where on the photograph—at the left, middle, or right?
8. Figure 2 shows two views of the same results. What is the relationship between the UV absorption bands
shown in Figure 2a and the line with two peaks in Figure 2b?
9. In their experiment, how did the investigators cause the bacteria to switch from incorporating N15 into
their DNA to growing with only light nitrogen isotopes, N14?
10. Based on the second paragraph on p. 676, how does the relative position of a band indicate how much of
the DNA was labeled with N15?
2
11. (a) From the legend of Figure 3, what was the average bacterial generation time (in hours) for these
experiments? (b) About how long would it take for four generations? (c) In Figure 3, what was the
experimental change made at time 0?
12. Examine the results of Figure 4, in which higher density DNA is to the right. (a) What is the density
difference between the initial DNA (0 generations) and the DNA after 1.0 generations? (b) How did the
DNA density distribution after 1 generation compare to that after 2.5 generations? (c) DNA at high
density (the farthest right band) is present at generation times 0, 0.3, and 0.7. At what other generations
is the high-density DNA present?
13. The second point of the Discussion section (p. 676) states that each daughter molecule receives one
subunit from the parent DNA. What pattern in the results supports this statement?
14. In the Discussion section, the authors draw connections between their own density results and the
Watson-Crick proposal for a possible DNA duplication mechanism. In order for you to see the
connections clearly, draw a quick sketch of three figures shown in this paper that represent the DNA
after 1 generation: (a) the sketch from Figure 4, a tracing of the peak after 1.0 generation; (b) the
schematic representation of the first-generation daughter molecules from Figure 5; and (c) the firstgeneration daughter molecules from Figure 6 (illustration of mechanism proposed by Watson and
Crick).
15. Examine your sketches for (b) and (c) in the previous question. In each sketch, the two images are the
same. If they were different from each other, how would the image in sketch (a) be different?
16. Summarize the “structurally plausible hypothesis” of Watson and Crick for the duplication of the DNA
3
molecule, as explained on p. 677.
C. General Conclusions and Extensions of the Work
17. At the end of the paper, the authors state that the nitrogen of a DNA molecule is divided equally
between two subunits and each daughter molecule receives one of these. (a) Which component of a
DNA nucleotide contains nitrogen? (b) How could you label another part of the DNA and repeat these
tests with another component?
18. In the introduction, the authors point out that hypotheses about DNA replication make different
predictions about the distribution of parental atoms into daughter molecules. Today, we often refer to the
model presented here as a “semi-conservative” model for replication, because components of each
strand stay together (that is, the strand is conserved) but the two strands of the parent molecule do not
stay together. What would happen in the distribution of parental atoms into daughter molecules if DNA
replication followed a “conservative” model in which the entire parent DNA molecule was conserved?
19. The Meselson-Stahl experiments and this paper are considered important, groundbreaking studies in
molecular biology. Decades later, the experiments are still admired, and “the paper is still held aloft for
its clarity” [T. H. Davis, Meselson and Stahl: The art of DNA replication, PNAS 101:17895–17896
(2004).]. Some of the clarity of the centrifugation results was due to the fact that the DNA was
fragmented during handling (although the experimenters did not know it at the time). As quoted in Davis
(2004), Stahl said that pipetting DNA was like “throwing spaghetti over Niagara Falls.” When DNA
breaks into fragments, the sugar-phosphate backbone is broken, forming many short double helixes.
Why did the fragmented DNA still show the same pattern of nitrogen density banding?
20. Imagine that you were a member of this research team and involved in these experiments. What could be
a possible follow-up test that extends this work?
4