Case Teaching Notes for
“The Case of the Druid Dracula: Clicker Case Version”

Introduction / Background

This case study is a “clicker case.” It combines the use of student personal response systems (clickers) with case teaching methods and formats. The case is presented in class using a series of PowerPoint slides punctuated by questions (called “clicker questions”) that students respond to before moving on to the next slide. In this way, students work through the material to understand (and solve) the problem presented in the case. Specifically designed for use in large introductory science classes, the method integrates lecture material, case storylines, student discussion, (clicker) questions, the clarification of answers to those questions, more lecture, and data.

This case was modified from a previous case developed by Peggy Brickman and published by the National Center for Case Study Teaching in Science (see http://www.sciencecases.org/druid_dracula/druid_dracula.asp). It centers on an actual crime committed in Great Britain and featured on the BBC program Crimewatch in December 2001. The case introduces DNA structure and the mechanism cells use to copy their DNA, and relates these concepts to similar processes used by forensic scientists for DNA fingerprinting. It also introduces students to the technique of gel-electrophoresis and to basic probability determinations. In the case, students are asked to apply their understanding of DNA structure and replication to identify the likely perpetrator of the crime.

The case is designed for use in an introductory biology course either for science majors or non-majors. It could potentially be further modified for use in upper level classes as well. The case assumes that students have a basic understanding of macromolecules and cell structure.

Objectives

In this case, students learn:

• the basic structure of DNA, including that DNA molecules are composed of two chains.
• that the two nucleotide strands in a DNA chain:
  • are complementary to each other (A-T, G-C);
  • have distinct 5′ end and 3′ ends; and
  • have opposite polarity (anti-parallel).
• how DNA is replicated, including:
  • the major enzymes involved in replication and the functions of these enzymes;
  • that during replication, each DNA strand can serve as a template for production of a new complementary DNA strand; and
  • that scientists can mimic the mechanism to replication to copy particular pieces of DNA using the process of polymerase chain reaction (PCR).
• how DNA sequences can vary from one person to the next, including that:
  • short-tandem repeats (STRs) are stretches of repeated nucleotide sequences that vary between chromosomes;
  • the probability that a particular combination of STRs might exist in an individual just by chance can be estimated statistically; and
  • scientists can identify individuals by examining these differences in DNA sequence.

Misconceptions

• DNA strands do not have a distinct polarity.
• The techniques used by scientists to study DNA are unrelated to processes seen in living organisms when in reality these often either use or mimic strategies used by living cells.

Classroom Management / Blocks of Analysis

This case was designed with large classes (>200 students) of science or non-science majors in mind. However, it could also be used effectively with small and/or more advanced classes. The case is designed for one 75-minute class period. Clicker questions are interspersed throughout the case to evaluate students’ understanding of the material being covered. We often include small group discussions when we administer these questions.

Teaching the Case

Slide 1, Slide 2, and Slide 3: The first three slides in the PowerPoint presentation introduce the case. Slides 2 and 3 may be excluded and the background information presented orally by the instructor or via a handout if desired. An interesting aspect of the case is an indication of the occult introduced in Slide 2. However, a more realistic interpretation is provided in Slide 3.

Slide 3: In addition to the sneaker print, students will often point out the blood stains on the windowsill relatively quickly. Once pointed out, the students can be asked what information these blood stains can provide. We usually ask the students to consider how this blood could have been used when their parents were their age. Some students will quickly point out that blood type was examined. Ask the students what different blood types they are. (Most will be able to provide the four major blood types, A, B, AB, and O.) Can blood type be used to identify an individual?

Slide 4: Clicker Question 1 asks students what their blood type is. Many of the students will not know their blood type, but you should have a few students of each type. Tell the students to assume the blood left at the crime scene matches one of the more common types they indicated in their answers. Can the culprit be identified now? Why not? If blood type can’t be used to identify the culprit, what good is it? (It can be used to eliminate suspects, but that is usually not enough to solve a case.) Several students at this point will suggest it can be used for DNA.

Slide 5 (animated): Unlike blood typing, DNA fingerprinting can be used to potentially identify specific individuals. In addition, very little DNA is needed for DNA fingerprinting and can be obtained from almost anything that has cells left behind by a person, such as semen, a cigarette butt, a used water glass, an eyelash, urine, etc. In bloodstains, the DNA comes from white blood cells as red blood cells lack a nucleus and nuclear DNA.

Most students will associate DNA fingerprinting with criminal prosecutions. However it can be used for a wide variety of purposes including criminal defense (see the Innocence Project, online at http://www.innocenceproject.org/, founded to assist prisoners who could be proven innocent through DNA testing).

Slide 6 (animated): Most students will probably not be familiar with how investigators are able to use DNA to identify individuals. This is a good time to explain how every cell in a person contains the exact same DNA (with a small number of exceptions) and that most DNA is the same from person to person. However, differences do exist in small regions of DNA between people, and that no two people (other than identical twins) have the exact same DNA. Scientists can look at these small differences to try to identify individuals.

Slide 7 (animated): An example of one of the differences that can be seen between people is the amelogenin gene. This gene is found on both the X and the Y chromosome, but in different forms. The copy on the X is nine nucleotides shorter than the copy on the Y chromosome. By looking at the size of the amelogenin gene in the DNA of the blood stains left behind at the crime scene, investigators can determine if the blood was left behind by a male or female. At this point it might be good to ask the students if they had a copy of the amelogenin gene, could they tell how big it was? Pieces of DNA are too small to see by eye.

Slide 8 (animated): Introduces the students to the concept of gel electrophoresis. Gels are usually made from agarose, long chains of carbohydrate extracted from seaweed. When heated in water, agarose is a liquid and can be poured into a mold. As it cools, it solidifies (like jello) and forms a three dimensional, porous network of fibers. DNA has a net (-) electrical charge; if DNA fragments are placed at one end of the gel and exposed to an electric current, the fragments will migrate towards the positive electrode (cathode). Smaller fragments are able to fit through the pores more easily and migrate faster than larger fragments. Once the gel has run, the DNA is stained using a fluorescent dye such as ethidium bromide and the fragments show up as bands on the gel. If a sample containing DNA fragments of known size is run at the same time (lane S in the picture), it is possible to estimate the size of the DNA fragments in unknown samples (lanes 1 and 2 in the gel).

Slide 9: Clicker Question 2 asks students how big the DNA fragment indicated is. The band indicated by the arrow has migrated a little bit farther than the 600 base pair reference band so it is a little bit smaller, probably around 580 base pairs. More advanced classes can use this question to demonstrate how to prepare a standard curve to get more accurate estimations.

Slide 10: This slide is used to get students thinking a little bit about how to make DNA. The intensity of most dyes is directly proportional to the amount of material that is bound/stained. Because the band in lane 1 is brighter, some students may mistakenly believe that it contains a larger DNA fragment than the band in lane 2. However, because the fragment in lane 1 migrated further, it must be smaller than the fragment in lane 2. How is it possible then for the band in lane 1 to be brighter (i.e., has more DNA) than the DNA fragment in lane 2? The answer is that each band contains many identical copies of the same DNA fragment. For DNA to be visible by gel electrophoresis and staining, tens of thousands of copies of a DNA fragment must be present.

Ask the students to think back to the crime scene and the small blood stain left behind. Would there be many thousands of copies of the amelogenin gene (or any gene) present? Even worse, what about a lip print on a glass used by a suspect or on a cigarette butt? This is a good time to remind students that each cell contains just two copies of each chromosome and this would mean that the cell contains just two copies of each gene. Also, red blood cells don’t have chromosomes (these cells eject their nucleus when they mature) so only the much less abundant white blood cells can provide the necessary DNA. In many cases, a DNA sample left behind at a crime scene will not be enough for gel electrophoresis to work. What can the investigator do? Solution: make more copies of the DNA that they were able to collect.

Slide 11 (animated): How do you make more copies of a piece of DNA? To understand this, students will need to understand the structure of DNA. This slide covers nucleotide structure, a single DNA strand, a double stranded DNA molecule, and the complementary and anti-parallel nature of DNA.

Slide 12: Clicker Question 3 checks to see if the students understand basic DNA structure. This question also introduces the idea of how to copy a DNA molecule. Ask students how they know what the correct sequence was for the other DNA strand. Point out that cells use the same logic to copy DNA.

Slide 13 (animated): This slide walks the students through the general steps of replication.

Slide 14 and Slide 15: These slides walk students through some of the major enzymes that are involved in replication and could be excluded if the course does not go into this much depth. Animations are especially good for demonstrating this process for visually oriented students. Many publishers provide animations with their textbooks. Animations can also be found at:

• DNAI: http://www.dnai.org/a/index.html
• Dolan Learning Center: http://www.dnalc.org/home_alternate.html
• PBS: http://www.pbs.org/wgbh/aso/tryit/dna/#

Some important points to make at this point:

• DNA replication starts at multiple points along a chromosome. The DNA strands at these sites separate, creating a “replication bubble.” The two ends of the bubble (where the separated and hydrogen bonded strands are adjacent to each other) form “replication forks.” As replication continues, the replication forks move in opposite directions down the chromosome expanding the size of the replication bubble. Using multiple replication bubbles in this manner speeds up the process of replication. A single DNA polymerase enzyme can copy approximately 1000 nucleotides / second but even at this speed it would take several weeks to copy all of the DNA in a cell. A cell can actually copy its DNA within hours.
• DNA polymerase must have an existing 3′ end to start producing a complementary DNA strand. Primase can produce a short complementary RNA strand that provides this starting point.
• Primase and DNA polymerase can only synthesize a new nucleotide strand in the 5′ to 3′ direction (reads the template strand in the 3′ to 5′ direction). As the replication fork moves down the chromosome, this means that one DNA polymerase complex will continuously follow the fork, producing a long complementary DNA fragment (leading strand). The DNA polymerase complex on the other strand (lagging strand) will be moving away from the fork. As a result, replication must be re-initiated repeatedly as the replication fork moves down the chromosome. This results in the production of many small complementary DNA fragments.
• The RNA primers are removed and replaced by specialized DNA polymerase later in replication and the adjacent fragments will be joined together by ligase.

Slide 16: Clicker Question 4 checks to see if students understand the functions of the major enzymes involved in replication.

Slide 17 (animated): Point out to the students that they need to make more copies of the DNA left at the crime scene for gel electrophoresis. This can be done by mimicking natural DNA replication with the technique of Polymerase Chain Reaction (PCR). A key difference between natural replication and PCR is that natural replication copies all of the DNA in a cell while PCR copies specific, targeted fragments. This slide explains how PCR works.

1. DNA strands are separated using heat rather than helicase enzyme.
2. Synthetic DNA primers are used to provide a starting point for DNA polymerase. The primers are complementary to a specific sequence of DNA and, in this manner, can target a specific stretch of DNA for replication. Primers are allowed to anneal to the template DNA by lowering the temperature.
3. A special heat-stable DNA polymerase is used.
4. By successively raising and lowering the temperature, DNA replication can be automated and takes place in a short period of time. It is possible to have several replication (PCR) cycles producing tens of millions of copies of a specific DNA fragment in less than an hour.

A good animation of PCR can be found at the Dolan Learning center website: http://www.dnalc.org/home_alternate.html

Slide 18: Clicker Question 5 checks to see if the students understand the process of PCR.

Slide 19: Clicker Question 6 requires students to use their understanding of both DNA structure and replication. A correct primer needs to be complementary to the 3′ end of the template strand. The top strand could be read from right to left (3′ to 5′) while the bottom strand could be read from left to right. The correct answer would be E, as it would be complementary to the 3′ end of the bottom strand.

Slide 20 (animated): Most forensic labs today use automated equipment for DNA fingerprinting. This slide shows how data collected by such equipment compares to standard gels. Green peaks are for sample DNA fragments, brown are for molecular weight controls.

Slide 21: Clicker Question 7 Asks students to interpret the data to determine which sample came from a male. They will have to remember that the amelogenin gene on the X and Y chromosomes are different sizes. The correct answer will be A. Females, XX, would produce two similar sized fragments that would appear as just one band. Males, XY, would produce two different sized fragments that would appear as two bands.

Slide 22: Clicker Question 8 asks students if this information alone would be enough to convict someone. Most will readily say no as half of the population is male and would still be potential suspects.

Slide 23 (animated): This slide introduces the students to additional markers used in DNA fingerprinting, namely, short tandem repeats (STRs). STRs are short stretches of DNA that are repeated several times one after another in a region of chromosome. The exact number of times the sequence is repeated can vary from chromosome to chromosome and can be used to distinguish between individuals.

Slide 24: Different chromosomes have different STR regions composed of different repeating sequences. If enough STR regions are used, each person should be unique.

Slide 25: If STR fragments are amplified by PCR, the fragments that are produced will vary in size depending on how many repeats of a sequence were present on the chromosomes being examined. The figure in this slide shows the number of repeats and DNA fragment size for three different STR regions, THO1, TPOX, and CSF1PO.

Slide 26: Hypothetical DNA fingerprint analysis from the crime scene in Wales, including positive and negative PCR controls, DNA from the blood stain (Evidence Sample), DNA from the victim, and DNA from two potential suspects. Can the students identify which suspect matches the DNA left at the crime scene? (Suspect #1).

Slide 27: Clicker Question 9 introduces students to the accuracy of DNA fingerprinting. If just one STR, THO1, was used and a suspect and evidence sample matched with both 5 and 7 repeats, how likely is it that the match is due to just random chance? There are two ways this could happen. Each individual has two chromosomes. One chromosome could have 5 repeats and the other 7 repeats (5 and 7). Alternatively, the first chromosome could have seven repeats and the second five (7 and 5). The probability that an individual might have any combination of 5 and 7 repeats would be:

(1/200 × 1/6) + (1/6 × 1/200) = 2/1200 = 1/600.

This applies concepts of probabilities from genetics (multiplication and addition rules) that may not have been covered yet. The main point students should realize is that one STR is not enough to identify a specific individual. To do this, additional STRs must be used to reduce the likelihood that someone may match a DNA fingerprint by random chance. If enough STRs are used, the likelihood of a random match can be reduced to 1 in several tens of millions or more.

Slide 28: As indicated earlier, this case is based on a true story. Standard police work identified Matthew Hardman as a suspect. Preliminary DNA testing provided enough evidence to arrest Hardman on suspicion of murder. During the arrest, a knife was found in his coat pocket. Subsequent DNA testing revealed two sources of DNA on the knife. One set of DNA was from Hardman and one matched the victim. The possibility of a random match was one in 73 million.

Assessment

This case was developed as part of an NSF-sponsored grant (# DUE 0618570) to determine whether clicker cases such as this one produced greater learning than the traditional lecture approach. As part of that project, the clicker cases had questions that were asked of students both before and after the class in which the material was presented. The questions were also used again during the final exam. These additional pre- and post-case questions are presented in the Answer Key.

Answer Key

Answers to the questions posed in the case study are provided in a separate answer key to the case. Those answers are password-protected. To access the answers for this case, go to the key. You will be prompted for a username and password. If you have not yet registered with us, you can see whether you are eligible for an account by reviewing our password policy and then apply online or write to answerkey@sciencecases.org.

Resources

Cellmark Diagnostics, Allele Frequencies for US Populations. Last accessed June 11, 2008.

http://dna-view.com/cellmark.htm

Forensic Science Service, Case Files: Mabel Leyshon—The Hunt for a Vampire. Last accessed June 11, 2008.

http://www.forensic.gov.uk/forensic_t/inside/news/list_casefiles.php?case=13

Kline, M.C., Redman, J.W., and J.M. Butler. 2001. “Training on STR Typing Using Commercial Kits and ABI 310/3100.” National Institute of Standards and Technology Website. Last accessed June 11, 2008.

http://www.cstl.nist.gov/div831/strbase/training.htm

Online Mendelian Inheritance in Man. Last accessed June 8, 2006.

http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=300391

Source for amelogenin information.

Slide Credits

Acknowledgements: This material is based upon work supported by the NSF Grant No. DUE-0618570. Any opinions, findings, conclusions, or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of NSF.

Date Posted: January 13, 2009.

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