1
The Case of the Druid Dracula: Clicker Case Version
Adapted from Brickman (2004). The Case of the Druid Dracula. National Center for Case Study Teaching in Science, University at Buffalo, State University of New York.
by Norris Armstrong, Terry Platt, and Peggy Brickman
2
CQ1: What is your blood type?
A: AB: BC: ABD: OE: Don’t know
3
Evidence in the Courtroom
• Primarily rape cases• Paternity testing• Historical/missing persons investigations• Military “dog tag”• Convicted felon databases
• Blood was previously used for blood typing• Now it is used as a source of DNA
Which cells in the blood contain DNA?
Uses for DNA fingerprinting
4
The Crime
In a quiet corner of Wales in the village of Llanfairpwll, 90-year-old Mabel Leyshorn was murdered. Her murder had been not only brutal (her heart had been hacked out), but also creepy. It appeared as if Mabel’s blood had been collected in a small kitchen saucepan and tasted. The murder showed other signs of the occult: a candlestick and a pair of crossed pokers had been arranged near the body.
- from BBC’s Crimewatch December 2001
5
The Crime SceneFurther investigation indicated that this was no supernatural villain at work: the murderer had worn tennis shoes which had left distinctive footprints under the glass door that had been shattered by a piece of broken garden slate. Moreover, the windowsill had bloodstains on it; with any luck, the evidence recovery unit hoped to use it to help determine who had committed the crime.
6
•Antiparallel
•Complementary
DNA Structure- Review
Two DNA chains
?′ end
3′ end 5′ end
3′ end
7
Copying DNA (Replication)
ATCGGACT
TAGCCTGA
--------
DNA strands are separated
Each single strand is used as a template to make a complementary strand
Two identical DNA molecules are produced
ATCGGACT
TAGCCTGA
- GA-T-
T -C -A -
ATCGGACT
--------
TAGCCTGA
--------
8
Enzymes Perform Replication• Helicases unwind DNA double helix.• Single Stranded Binding Proteins hold separated
DNA strands apart.• Primase makes a starting point (primer).• DNA polymerase connects new complementary
bases.• Ligase attaches pieces together.
CQ2: Which enzyme elongates the new DNA daughter strand?A.DNA polymerase IB.DNA Polymerase III
9
Enzymes Perform Replication
Replication fork
10
DNA in the CellDNA in the Cell
Target GeneTarget Gene
chromosome
double stranded DNA molecule
individual nucleotides
11
Example: Amelogenin Gene • Target sequence to be copied in this gene is located in Intron 1• Gene function: tooth enamel development • Copies on both X and Y chromosome• X copy is different from Y copy
X copy is shorter than Y copy
Y:
X:
--- indicates missing bases on X
5’CCCTAGGGTCTA---------GTGTGTTGATTC 5’3’GGGATCCCAGAT---------CACACAACTAAG 3’
GTGTGTTGATTC 3’CACACAACTAAG 5’
5’CCCTAGGGTCTATAACGCCTAGTGTGTTGATTC 5’3’GGGATCCCAGATATTGCGGATCACACAACTAAG 3’
12
CQ3: Below is one strand from part of the amelogenin gene. What is the nucleotide sequence of the complementary strand?
A: 3′ -ACTGTTAGATT-5′ B: 3′ -GGGACCCGAGA-5′ C: 5′ -GGGACCCGAGA-3′ D: 3′ -CCCTGGGCTCT-5′ E: 5′ -CCCTGGGCTCT-3′
5’-CCCTGGGCTCT-3’
13
CQ4: How would DNA replication be affected if ligase were not available?
A: The template strands would not be able to separate.
B: Replication would result in many small segments of DNA instead of a complete molecule.
C: Complementary RNA would be produced but not complementary DNA.
D: The DNA strands would separate but replication would not be able to start.
E: The DNA strands produced by replication would not be complementary to the template strands.
14In 32 cycles at 100% efficiency, 1.07 billion copies of targeted DNA region are created
In 32 cycles at 100% efficiency, 1.07 billion copies of targeted DNA region are created
Amplifying DNA with PCR (Polymerase Chain Reaction)Amplifying DNA with PCR (Polymerase Chain Reaction)
Target region
Thermal cycleThermal cycleThermal cycle
15
CQ5: In order to conduct forensic tests, you need many copies of the amelogenin gene target sequence, which you will make using Polymerase Chain Reaction (PCR). PCR follows the steps of replication. Which of the following would allow replication to begin?
A: Add short stretches of single stranded DNA complementary to the sequence at either end of the gene.
B: Add DNA polymerase enzyme.
C: Break the covalent bonds that hold the double helix together.
D: Break the hydrogen bonds that hold the double helix together. (denature)
16
CQ6: Which of the strands of DNA could act as one of the two primers for the DNA sequence shown below?
5’-CCCTGGGCTCTGTAAATGTTTCTAAGTG-3’3’-GGGACCCGAGACATTTACAAAGATTCAC-5’
A: 3′ -ACTGTTAGA-5′
B: 3′ -AAATTTGGC-5′
C: 3′ -ATGCTTTGA-5′
D: 5′ -GGGACCCGA-3′
E: 5′ -CCCTGGGCT-3′
17
CQ7: If both of these were run through gel electrophoresis, which segment would go farther through the gel?A.YB.X
Y:
X: 5’CCCTAGGGTCTA---------GTGTGTTGATTC 5’3’GGGATCCCAGAT---------CACACAACTAAG 3’
GTGTGTTGATTC 3’CACACAACTAAG 5’
5’CCCTAGGGTCTATAACGCCTAGTGTGTTGATTC 5’3’GGGATCCCAGATATTGCGGATCACACAACTAAG 3’
18
Gel Electrophoresis: Sizing DNA Fragments
(-) Negative electrode
(+) Positive electrode
#bp?
#bp?
19
CQ8: The DNA fragment indicated is approximately ____ base pairs in size.
bp?
A: 300B: 350C: 580D: 600E: 700
20
Why do the two DNA fragments indicated differ in how bright they appear?
21
Additional Markers
---TCAT------TCAT---
Short Tandem Repeats (STRs)Short Tandem Repeats (STRs)
•Same pair in suspect 2:
•Different people have different numbers of repeats on their chromosomes
•Chromosomes 11 of suspect 1:
22
Positions of other STR
regions
CSF1PO
TH01
TPOX
AMEL
Each person is unique
23
Druid Dracula: DNA testing
• With kits, just add DNA sample with primers for amelogenin (XY) & different STR regions.
• Amplify and run on gel (electrophoresis).• An allele ladder shows all varieties in a population.
24
Automated gels
MW Amelog.
101 bp110 bp
25
CQ9: The blood left at the crime scene was from a male. Which of the following DNA profiles could have come from the suspect?
A:
B:
26
CQ10: Is this enough to convict a suspect?
A: Yes
B: No
27
CQ11: Which suspect presents a profile compatible to the sample found at the crime scene?A.Suspect 1B.Suspect 2
What are the Odds?• There are only a few different numbers of repeats that
are seen in our population.– Ex: only 5 different TPOX STRS
• After testing thousands of DNA samples, researches know the distribution of these different STRS in the general population.
28
THO1 TPOX CSF1PO
5: 1/200 8: ½ 9: 1/40
6: ¼ 9: 1/8 10: 1/5
7: 1/6 10: 1/18 11: 1/3
8: 1/7 11: 1/5 12: 1/3
9: 1/6 12: 1/20 13: 1/10
9.3: 1/3 14: 1/50
10: 1/100
29
CQ12: What is the probability that two people might have 5 and 7 repeats for the STR THO1 by random chance?
STR THO1 allele frequencies
5 6789
9.310
1/2001/41/61/71/61/3
1/100
A: 1/200B: 1/206C: 1/600D: 1/1200E: 1/2600
What are the Odds?CQ13: What is the probability that someone else at random would have the same pattern of THO1 5 & 7, TPOX 8 & 9, and CSF1PO 11 & 12?
A.1/1600
B.1/7200
C.1/17600
D.1/172800
E.1/1555200
30
THO1 TPOX CSF1PO
5: 1/200 8: ½ 9: 1/40
6: ¼ 9: 1/8 10: 1/5
7: 1/6 10: 1/18 11: 1/3
8: 1/7 11: 1/5 12: 1/3
9: 1/6 12: 1/20 13: 1/10
9.3: 1/3 14: 1/50
10: 1/100
31
Hardman’s Arrest• 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 from Hardman and one matching the victim. The possibility of a random match was one in 73 million.
• A search of Hardman’s dwelling produced magazines and evidence of accessing internet sites featuring how to become a vampire.
• Matthew Hardman was found guilty of murder on August 2, 2002, and sentenced to life imprisonment.