Discovering the Structure of DNA

Instructions: Read through the clues in each set below. As you do, respond to questions in the text.

For most of human history, we have had an understanding that children inherit traits from their parents. Ancient humans used this observation to selectively breed plants for desired traits–they understood that corn plants with larger kernels would produce offspring that also had larger kernels. In the 17th & 18th centuries, as science began to mature as a rigorous pursuit of knowledge, people began more specifically experimenting with–and observing–the movement of traits from one generation to the next.

In 1866, an Austrian monk by the name of Gregor Mendel had collected an enormous body of data on the inheritance patterns of various traits in peas. From the data, he concluded that some type of units were responsible for the passage of traits from one generation to the next.

What is perhaps most fascinating about this particular bit of science history is that no one, not even Mendel, had any idea about the mechanism of inheritance. A phenomenon had been extraordinarily well-documented–traits can move from one generation to the next–but no one could satisfactorily explain why or how. Mendel used the word gene in his published research, but he did not know what a gene actually was. He just needed some word to use to describe the “whatever-it-was” that was being transferred from parent to offspring. Gene was not originally a structural term, but a functional one. It comes from the Greek word genesis, roughly meaning “race” or “offspring.” Something is being handed off from parent to offspring, Mendel knew, and he called it a gene. However, he offered no suggestions about the physical nature of the gene. This set the stage for what became the driving question of biological investigation for the next century: what actually is the gene?

Clues Set 1

1. In 1868 Friedrich Miescher, a swiss scientist working at the University of Tubingen in Germany. was looking at pus found in surgical bandages when he discovered that when pepsin (an enzyme known to break apart proteins) was added to chromosomes, atoms of oxygen, carbon, hydrogen, and nitrogen were detected. This made sense, because these were the atoms known to be present in proteins. But he also detected phosphorus atoms. Because phosphorus atoms are never in proteins, he suspected that another type of molecule, in addition to protein, must be present in chromosomes. He named the new molecule nuclein. Twenty three years later he suggested that nuclein could be a molecule behind heredity. Nuclein is now known as DNA.

left11430002. Acetabularia is a single celled marine algae. Despite this organism’s single-celled status, it grows large enough to be easily visible to the naked eye. One species, A. mediterranea, has a smooth cap, while another species, A. crenulata, has a more ragged cap (see diagram below). In all species of Acetabularia, the nucleus is located in the bottom of the stalk.

The diagrams at the left show the outcome of nuclear transplantation experiments. Note what happens when the nucleus of A. crenulata is grafted on to the stalk of A. mediterranea.

What does this say about the role of the nucleus in determining the traits of the cell?

3. During the 1880s, German biologist Walter Flemming studied the behavior of chromosomes in reproducing cells. His work showed that new individuals begin with the union of sperm and egg cells. The sperm cells contain chromosomes and often little else, while eggs contain chromosomes and other cytoplasmic structures.

What does this establish about the role of chromosomes?

4. During the early 1900s, German chemist Robert Feulgen discovered that all body cells of any particular organism contain precisely the same amount of nuclein but that the amount of protein varies from cell to cell. He also found that egg and sperm cells contain exactly one-half the amount of nuclein present in body cells.

Why is that significant?

-25400741045005. In the 1920s a physician turned chemist Phoebus Levene was the first to discover the order of the three major components of a single nucleotide; the first to discover the sugar ribose was in RNA; the first to discover deoxyribose was in DNA; and the first to correctly identify the way the RNA and DNA are put together in the order phosphate-sugar-base to form units. He called each of these units a nucletide, and stated that the DNA molecule consisted of a string of nucleotide units linked together through the phosphate groups, which are the ‘backbone’ of the molecule. His ideas about the structure of DNA were wrong; he thought there were only four nucleotides per molecule. He even declared that it could not store the genetic code because it was chemically far too simple. However, his work was a key basis for the later work that determined the structure of DNA.

A representation of a nucleotide containing the nitrogenous base adenine is shown to the left.

6. During the late 1940s, English scientists discovered that DNA (like sugar) can crystalize when water is removed. This fact suggested that the atoms in DNA must be arranged in a very orderly way, perhaps with many repetitions of a fairly simple pattern. Protein, by contrast, doesn’t crystalize.

What was this genetic substance that passed on information, determined an offspring’s visible traits, and also accounted for the incredible diversity to be found among all living things? Some scientists thought it must be a protein. They knew that proteins are present in large quantities in the cell and that they carry on numerous functions. Proteins are made of 20 different subunits called amino acids. These subunits can be joined in a great variety of combinations. Supporters of proteins as the molecule for heredity thought that this variety would allow for the diversity we see in organisms. They imagined this in much the same way as the 26 letters of the English alphabet. These letters can be placed in a variety of ways to produce an immense quantity of words.

Other scientists thought that DNA was the molecule of heredity. They noted that large amounts of DNA were also present in cells (nearly 2 meters worth). And like the suspect of a murder, it kept turning up in experiments. However it seemed too simple a molecule. It only has six subunits: deoxyribose (a sugar), phosphate and four nitrogenous bases (A, G,T, C). The debate ranged hot and heavy. Which was the molecule of heredity?

Skim Clues 1 – 6. Summarize these clues down to about 4 or 5 bullet points and write them below.

Clues Set 2

522541550807. In 1908 Thomas Hunt Morgan and his students, through experiments with fruit fly mutations discovered that genes were carried on chromosomes. Morgan had become interested in species variation, and in 1911, he established the “Fly Room” at Columbia to determine how a species changed over time. For the next 17 years, in a 16 X 23 ft. room, described by many as cramped, dusty, smelly and cockroach ridden, Morgan and his students did ground-breaking genetic research using Drosophila melanogaster, fruit flies. Though initially against the idea that the behavior of chromosomes can explain inheritance, Morgan became the leading supporter of the idea. Morgan and his students developed the ideas, and provided the proof for the chromosomal theory of heredity, genetic linkage, chromosomal crossing over and non-disjunction. Because of Morgan’s dramatic success with Drosophila, many other labs throughout the world took up fruit fly genetics. He received the Nobel Prize in 1933

How did Morgan’s work influence the debate over DNA or Protein being the Molecule of Heredity?

left688975008. During the influenza Pandemic of 1918-1919, many of the deaths associated with the flu virus were actually caused by secondary infections involving a type of bacteria called Pneumococcus. Infection with this bacteria led to pneumonia, a lung inflammation in which the air sacs fill with pus. This made understanding the biology of Pneumococcus an important research topic in the 1920s. One of the researchers whose focus was Pneumococcus was the British bacteriologist Frederick Griffith, who was studying Pneumococcus in the hopes of developing a vaccine against pneumococcal infection.

Pneumococci bacteria come in two varieties, or strains. In one strain, the pneumococci are surrounded by a capsule, a layer of polysaccharide just outside of the cell membrane. These are called “smooth.” In the second strain, the cells lack a capsule. These are called “rough.” You can see the difference between these two strains in the picture of bacterial colonies to your left.

These two strains vary in terms of virulence (ability to cause disease), as you can see in the upper part (1 and 2) of the diagram to your left.

40195500In 1928, Griffith carried out a series of experiments on Pneumococcus, which are summarized in the diagram and the text that follows.

In diagram 1, smooth strain bacteria were injected into mice. The mice developed pneumonia and died.

In diagram 2, rough strain bacteria were injected into mice. The mice survived.

In diagram 3, Griffith heated smooth strain bacteria, which killed the cells. When he injected these heat-killed smooth strain bacteria into mice, the mice lived.

Next, Griffith took these heat killed, smooth-strain bacteria and mixed them with living rough strain bacteria (the mixture is shown at 4). He then injected this mixture into mice. The mice died.

When blood was drawn from these dead mice, Griffith observed living smooth strain bacteria (shown at 5).

What can you conclude from this series of experiments? Be sure to write down your conclusion in the space below before proceeding to the next page.

It didn’t take long for the scientific community to suspect that Griffith’s transforming principle was probably related to Mendel’s gene. The challenge of determining its physical identity remained

8. The result of Griffith’s work was identification of an unknown “transforming factor.” Something in heat-killed, smooth-strain Pneumococcus bacteria could transform non-virulent rough-strain bacteria into the virulent smooth-strain bacteria. But what was the chemical nature of this transforming factor?

In 1944, after many years of chemical analysis and experimentation, It fell upon Oswald Avery, working with his colleagues Colin MacLeod and Maclyn McCarty at the Rockefeller University Hospital in New York City, to figure out what this transforming factor was. Avery’s experiments involved taking the transforming factor and digesting it with various enzymes. The diagrams below summarize what they did and what they found.

As is shown in “1,” they started with heat-killed, smooth strain bacteria (the virulent variety). Remember that even though the cells were dead, they still contained the “transforming factor.” -38099320934They broke down the cells, and removed the lipids and carbohydrates, leaving them with a liquid containing the possible contenders for the transforming factor: nucleic acids and protein. This liquid was in test tube “2.” Keep in mind that nucleic acids come in two varieties: DNA, and a closely related molecule, RNA.

They subjected the nucleic acid/protein-containing liquid to three enzymes. Enzyme A would digest the protein, creating a protein-free liquid. Enzyme B would digest RNA, and enzyme C would digest DNA. The adding of these enzymes is shown in “3”.

Then they added non-virulent, rough strain bacteria, and allowed the bacteria to grow (shown in row “4”).

The results are shown in “5.” In the flasks where the protein and the RNA had been digested (columns “A” and “B”), they found both rough and smooth strain bacteria. In the flask where DNA had been digested, there were only rough bacteria.

In the space below, describe what Avery and his colleagues had established, and how they had established it.

9. In the early 1950s, Alfred Hershey and Martha Chase, from the Cold Spring Harbor Laboratory in New York, were working with viruses. The particular virus they worked with is called a phage: a virus that attacks bacteria. Here’s what Hershey and Chase knew about viruses as they were setting up what turned out to be a landmark experiment.

Phage viruses infect bacterial cells, turning them into virus factories.

Phage viruses are composed mostly of DNA and protein.

Phage have a structure that consists of an outer coat, and an inner core. When phage infect bacterial cells, they leave their coat outside the cell and inject something inside which initiates the cycle of viral takeover.

Hershey & Chase knew the following information about the chemical elements in DNA and protein:

Carbon Oxygen Hydrogen Nitrogen Sulfur Phosphorous

Found in DNA? X X X X X

Found in Protein? X X X X X They knew that if they could track the movement of sulfur, they would have information about the movement of protein. If they could track the movement of phosphorous, they would have information about the movement of DNA. However, as you may know, atoms are tiny! Much too tiny to see with microscopes. And protein and DNA are both colorless in solution, even in large quantities. Hershey & Chase needed a tool for visualizing these invisible atoms. They used radioactive labeling, which is a tremendously useful and still commonly used method in modern biology. They did this by growing bacteria on two separate food sources. In group one, they used a radioactive isotope of phosphorus, 32P. In the second group, they used a radioactive isotope of sulfur, 35S. Next, they infected these bacteria with phage. When phage take over bacterial cells, they use them as virus factories to assemble more phage. As a result, whatever the bacteria are made of, the phage will be made of (because even for viruses, “you are what you eat.”). Consequently, the phage that would be produced in group one (with radioactive phosphorus) would have radioactive phosphorus built into their DNA. By contrast, the phage produced in group two would have radioactive sulfur built into their protein.

451485074930With this labeling system, all that Hershey and Chase had to do was to allow phage to start their infection cycle. If they could isolate infected cells just at the moment after the phage had injected their genes inside the cells that they were attacking, they could then test to see what the phage had injected inside: protein, or DNA. Here’s how they did it.

In row 1, at the top of the diagram to the right, you can see the radioactively labeled phage. These phage are allowed to infect cells that have been grown on normal, non-radioactive growth media (shown in row 2). That way, if any radioactivity appeared in the infected cells, Hershey and Chase would know that it was brought in by the radioactive phage.

Now, remember that the goal was to figure out what the genetic material that the phage were injecting into the cells was made of. Hershey and Chase didn’t want the phage to go through their entire reproductive cycle, because that would make interpretation of the results impossible (see if you can figure out why after you read what follows).

So after just enough time passed for the phage to inject their genes into their bacterial victims, Hershey right59690and Chase placed the mixture of phage and bacteria into a blender (shown in row 3). The blender shook the attacking phage off of the cells they had just attacked (row 4). Then they broke the cells open (using chemicals that dissolve the cells’ membranes while not destroying protein or DNA) and placed the mixtures in a centrifuge. A centrifuge spins test tubes around at high speed, and, as it does, it separates whatever is in the test tube by density. The result was a test tube with liquid on top (called the “supernatant”) and a pellet of denser, cellular debris on the bottom (see row 5).

Note the results: In the treatment with radioactive phosphorus, 35P was found in the pellet of cellular debris at the bottom of the test tube. And in the treatment with radioactive sulfur, 32S was found only in the supernatant (and not in the debris pellet).

How would you interpret this? Write down your interpretation below before continuing.

Clues Set 3 (1952)

464820082550010. At this point, it was clear that genes were made of DNA. So what structure could DNA have that could allow it to store and transmit genetic information? A key clue related to DNA’s structure was provided by the work of Rosalind Franklin. Franklin, working with her colleague Maurice Wilkins and her graduate student Raymond Gosling at Cambridge University in England was trying to decipher the structure of DNA by crystallizing it, and then photographing the crystals using X-rays. The X-rays would create a diffraction pattern that could be used to infer the arrangement of the atoms in the DNA crystal. Some of the thinking involved in interpreting these images is demonstrated below by Stephen Carr:

“…The X-ray crystallograph (above, also known as “Photo 51”) shows an exceptionally clear diffraction pattern of a crystallized DNA molecule. The X-pattern in the middle is characteristic of a helical molecule with regular repeats.”

The image also allowed one to infer the distance between the monomers making up the DNA polymer (3.4 Angstroms, or 0.34 nanometers), and that the width of the DNA stayed constant throughout its entire length.”

Historians of science have speculated that Franklin and Wilkins, with a bit more time, might have deduced the structure of DNA. But the prize was won by two other researchers who were also working in Cambridge. Novices James Watson, and Francis Crick would prove the first to discover the structure of the DNA molecule. Watson was an American prodigy who attended to University of Chicago at 15 and a PhD from Cambridge by 22. He met the older Crick in Cambridge and together assumed that if you could determine the shape of DNA you could determine how it did what it did. They weren’t actually assigned to work on DNA and at one point were ordered to stop. They did not carry out laboratory experiments themselves but based their hypothesis on data accumulated from others. Franklin was constantly being bullied by her supervisor Maurice Wilkins and Watson and Crick. She started making false comments about her work in an effort to try to protect it. Finally in January of 1953 Wilkins stole the X-ray from her and gave it to Watson and Crick. Watson and Crick also had access to other clues (which others had access to as well):

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11. Scientists now knew that DNA functioned as the storage unit and transmitter of genetic information. How were those six subunits arranged in the molecule so that DNA could be the bearer of vast amounts of information and code for incredible diversity seen in all living things? Erwin Chargraff added an important piece to the puzzle with his experiments. He showed that the proportions of nitrogenous bases found in DNA were the same in every cell of an organism in a given species. But he showed that the proportions varied from species to species. He also provided an important piece of data about the ratios of nitrogenous bases.

12. Additional evidence indicated that the DNA helix contains strands of alternating phosphate groups and sugar molecules linked by covalent bonds.

13. In the spring of 1953, Watson arrived at the lab early one day, cut out cardboard models of nucleotides containing adenine, thymine, guanine, and cytosine molecules, and began arranging them in various combinations and patterns on his desk. He discovered that an adenine-thymine pair presumably held together by relatively weak hydrogen bonds is identical in shape to a guanine-cytosine pair also held together by hydrogen bonds.

Write a summary of what you learned above from clues set 3

Of all of the research and discoveries into the structure of DNA who’s work do you believe was the most valuable and why?

DRAW A CONCLUSION:

From this information, Watson and Crick were able to build a model that satisfied all the known data. They knew that the molecular structure must be varied, carry a large amount of information, replicate inself before cell division and code for traits. Their paper, published in April 1953, set off great excitement in the biological community, initiating the explosion of DNA research that continues today. Watson, Crick and Maurice Wilkins were awarded the Nobel Prize in Medicine in 1962. Rosalind Franklin did not share in the Noble Prize as she died in 1958 of cancer which most likely developed as a result of her work with X-rays.

Now it’s your turn to discover the same secret:

YOUR FINAL JOB: After you’ve drawn inferences from all the clues above, your job is to create your own model of DNA. Take a photograph of your DNA model, and insert it below.

PLACE PHOTO OF YOUR DNA MODEL HERE.

OPTION 1: Print out 3 the last 2 pages of this handout. Cut out the shapes, and then follow the instructions below.

1) Create a model of DNA that contains at least 9 nucleotides long

2) On your model, indicate which bonds are covalent bonds, and which bonds are hydrogen bonds.

3) Take a photo and insert it above.

4) Create a bullet point list where you explain how the model you’ve created is supported by various pieces of evidence that you’ve gleaned from this activity.

5) Propose a method by which DNA could be replicated (something that Watson and Crick did in their 1953 paper describing the structure of DNA).

OPTION 2: If you don’t have a printer, grab a pencil, and using the shapes on the last page as a guide, draw a model of DNA that’s at least 9 nucleotides long.

OPTION 3: Use the PowerPoint File in Schoology. Use the pieces inside to create your own model of DNA, which you can take a screen shot of and then paste it above.