Student Case Study
The following is a case study that is designed to support you in answering the question ‘How does DNA replicate?’ It is assumed that you already know the structure of DNA and you now want to find out how DNA makes copies of itself during cell division.
The case study is designed to provide you with an opportunity to find out the answer to the question yourself, using the real experimental data from the scientists who first successfully answered it in 1958.
It is intended that by completing this case study you should not only learn the mechanism by which DNA replicates, but should also have had the opportunity to develop the scientific enquiry skills.
Higher Biology
1. b. (i) DNA can direct its own replication.

 http://www.sumanasinc.com/webcontent/animations/content/meselson.html
http://highered.mcgrawhill.com/olcweb/cgi/pluginpop.cgi?it=swf::535::535::/sites/dl/free/007243
7316/120076/bio22.swf::Meselson%20and%20Stahl%20Experiment
 http://www.learnerstv.com/animation/animation.php?ani=20&cat=biology
 http://www.dnalc.org/view/15331 -Proposed-models-of-DNA-replication-
Matthew-Meselson-.html
 http://www.dnalc.org/view/15880 -Models-of-DNA-replication.html
 http://www.dnalc.org/view/15879 -Semi-conservative-replication.html

HOW DOES DNA REPLICATE?
Activity 1
In April 1953 Francis Crick and James D.
Watson had a paper published in the Journal
Nature entitled ‘A Structure for Deoxyribose
Nucleic Acids’.
In this paper they wrote:
‘We wish to put forward a radically different structure for the salt of deoxyribose nucleic acid. This structure has two helical chains each coiled around the same axis.’
Watson and Crick (1953)
Watson & Crick
Image from Preoccupation
Use this information and your prior learning to draw an annota ted diagram of Watson and Crick’s structure of DNA.
Activity 2
So, we know the structure of DNA – but why does it replicate?
Why does DNA need to be able to copy itself?
Activity 3
DNA replication occurs as part of a process that occurs in nearly every cell in our bodies.
Name this process and summarise it as a diagram in your jotter.

Student Case Study
Activity 4
In the same 1953 paper, Watson and Crick went on to say:
‘If an adenine forms one member of a pair, on either chain, then…the other member must be thymine; similarly for guanine and cytosine…It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material. ’
Watson and Crick (1953)
DNA can direct its own replication, but how does this happen? Watson and
Crick hinted at a possible mechanism, but they were not able to supply any experimental evidence to support their theory. What do you think? How does one DNA molecule turn into two?
Perhaps an entire new DNA molecule is made from an ‘old’ DNA molecule , like this:
DNA
R EPLICATION
In this hypothesis, the two new DNA strands occur together in one daughter cell, the two old in the other.
Conservative model
‘old’ DNA ‘old’ DNA ‘new’ DNA
There are at least two other possible hypotheses to expla in how new DNA strands might be made. Draw these in your jotter. Once you have drawn the other models, describe them and try to find out their names.

HOW DOES DNA REPLICATE?
Activity 5
Matthew Meselson and Franklin W. Stahl set out to answer this very question. In 1958 they published a paper that outlined their experimental procedure for determining the mechanism of DNA replication. They introduce the paper by saying:
‘Hypotheses for the mechanism of DNA replication differ in the predictions they
Meselson & Stahl
Image from achievement.org
make concerning the distribution among progeny molecules of atoms derived from parental molecules. Radioisotopic labels have been employed in experiments bearing on the distribution of parental atoms among progeny molecules in several organisms. We anticipated that a label which imparts to the DNA molecule an increased density might permit an analysis of this distribution by sedimentation techniques.’
Meselson and Stahl (1958)
What does this mean? Look up any words you do not understand and explain in your own words what Meselson and Stahl are saying in the above quote.
Activity 6
Meselson and Stahl’s experiment relied on the different properties of the two isotopes of nitrogen, 14 N and 15 N. Isotopes can be distinguished on the basis of their mass: 14 N is lighter than 15 N; neither of these isotopes is radioactive.
Why is it important that these isotopes are not radioacti ve?
Activity 7
Meselson and Stahl grew bacteria in a medium where their nitrogen source, used to synthesise nucleotides and eventually DNA, was the ammonium ion
(NH
4
+ ) containing heavy 15 N. After several generations, all the DNA in these bacteria would be heavy.
Where are the nitrogen atoms in DNA?

Student Case Study
The parental bacteria were then placed in a fresh medium containing light
( 14 N) ammonium ions and allowed to divide and copy their DNA just once to produce generation 1 . A sample removed now from this beaker would contain generation 1 DNA.
Some of the generation 1 bacteria were then allowed to divide once more in
14 N medium to produce generation 2 bacteria.
Bacteria cultured in medium containing heavy nitrogen
( 15 N)
Bacteria transferred to medium containing light nitrogen
( 14 N)
DNA sample taken after 20 minutes.
Generation 1
DNA sample taken after 40 minutes.
Generation 2
To find out which form of replication is occurring, Meselson and Stahl isolated and separated the DNA from generation 1 and generation 2. This was done by density gradient centrifugation. This technique allows the separation of molecules on the basis of their density.
After centrifugation, the tubes were irradiated with ultraviolet light. DNA absorbs this light so the position of the DNA in the centrifuge tube shows up as a black band.
Light 14 N DNA is found at the top of the centrifuge tube and heavy 15 N DNA is located at the bottom of the tube. Any mixture of 14 N and 15 N DNA is found in the centre of the tube. light 14 N DNA
14 N and 15 N hybrid DNA heavy 15 N DNA

HOW DOES DNA REPLICATE?
Before you find out which of the models described above is the correct one, you must first work out what actual results Meselson and Stahl would have obtained for each of the predicted models.
Activity 8
In the table below, draw the expected centrifuge tube for each of the predictions for generation 1 and generation 2. The centrifuge tubes for the conservative model have been done for you.
Prediction
Model Conservative
Semiconservative
Dispersive
Generation 1
Generation 2
Activity 9
On the following page you will find the actual results from Meselson and
Stahl’s experiments. On their diagram the heavier DNA is found on the right.
They conducted a range of experiments. You will need to identify which of two results show generations 1 and 2. Compare Meselson and Stahl’s results with your own predictions above to determine which of the models of DNA replication their experiment supports.

Student Case Study

HOW DOES DNA REPLICATE?
Activity 10
Which is the correct model of DNA replication?
Explain how you arrived at this conclusion.
You now need to demonstrate your understanding of how DNA replicates and take your understanding further.
Your task is to prepare a resource that could be used by students learning about DNA replication using the following as your title:
How does DNA replicate?
It is up to you to choose how to present your understanding but through your resource you must:
 demonstrate a clear understanding of semi -conservative DNA replication
 explain why the data from Meselson and Stahl’s experiment supports semiconservative DNA replication over othe r hypotheses
 outline the steps involved in semi -conservative DNA replication.
You should consider the following questions when preparing your resource:

Why and when does DNA replicate?

What is the model of DNA replication?

What evidence supports this model of DNA replication?

How does the structure of DNA relate to the mechanism of replication?

Which enzymes catalyse DNA replication?

What is the sequence of steps involved in DNA replication?
Use your jotter to plan your resource.

Student Case Study
Now that you know how the model of DNA replication was tested, what about the enzyme which catalyses DNA replication – DNA polymerase ? What do we know about this enzyme? Who discovered it? When did they discover it?
What methods did they use?
Find out what you can about the discovery of DNA polymerase and summarise your findings.
Useful weblinks
 http://www.jbc.org/content/278/37/34733.full?maxtoshow=&HITS=10&hit s=10&RESULTFORMAT=&searchid=1&FIRSTINDEX=30&resourcetype
=HWCIT
 http://www.dnalc.org/resources/nobel/kornberg_ a.html