DNA Fingerprinting

1
DNA Fingerprinting
Ray McGovern Nabil Hafez John Leonard
November 29, 2004
2
Overview

• DNA Fingerprinting developed by Sir Alec J.
  Jeffreys, at the University of Leicester, UK.
• Leicestershire, saw the first exoneration and
  conviction bases of DNA evidence, in the late
  1980s.
• We will cover
• What is DNA Fingerprinting?
• Courts Cases
• Probabilities associated with DNA fingerprinting

3
Genetics Basics

• 100 trillion cells
• 23 pairs of chromosomes
• 99.9 identical between humans
• 0.1 or 3 million bases accounts for variation
• Est. 20,000 25,000 genes
• Up to hundreds of alleles per gene (blue or brown
  eyes)

Courtesy DOE
4
Allelic Differences

• Repeats vary from individual to individual but
  are also inherited
• Homozygous vs Heterozygous

Courtesy http//www.howstuffworks.com
5
Polymorphism Variation

• VNTR (minisatellites)
• Variable number of tandem repeats
• Repeat unit up to 25 bp (unit 20 kb)
• ACAGGGTGTGGGGACAGGGTGTGGGG
• vs
• AGTAGTAGTAGTAGTAGT
• STR (microsatellites)
• Short tandem repeats
• repeat unit less than 13 bp (unit lt150 bp)

6
Where do we get the DNA?

• Blood - extracted dry or wet (WBC)
• Buccal (cheek cells) easy to obtain
• Semen dried can be analyzed years
  later
• Hair roots
• Saliva
• Skin cells

7
RFLP VNTR based

• Restriction Fragment Length Polymorphism

Courtesy http//www.howstuffworks.com
8
Autoradiograph
DNA evidence
Suspect 1
Courtesy http//www.howstuffworks.com
9
STR (PCR) Typing

• Use PCR (polymerase chain reaction) to amplify
  DNA
• Primer sequence
• from locus region
• (locus chromosomal
• location of genetic
• marker or repeat)
• Obtain Product
• Run Gel

Courtesy Fan Sozzi
10
STR multiplexing

• Analyzed by computer and analyst
• This one is 8 STR loci on one gel
• 1 in 100,000,000 discrimination

Courtesy Genelex
11
RFLP vs STR typing

• RFLP
• Older method (80s)
• Less sensitive suitable for blood, requires
  large amount of high quality DNA
• Slow 7-10 days, labor intensive, one gene
  analyzed at a time
• Fewer indvidual tests needed, genes examined have
  gt100 alleles high power of discrimation

• STR (PCR)
• Newer method (90s)
• More sensitive can use degraded DNA from minute
  sources
• Fast 2-6 days, high throughput, simultaneous
  analysis of many genes
• More individual tests needed, genes examined have
  between 10-15 alles, low power of discrimination

12
CODIS(just like on CSI)

• Combined DNA Index System
• 1997 - National FBI Database composed of 13 STR
  Locis
• Probability that DNA from unrelated individuals
  would generate the same 13 STR profile would be
  less than 1 in a trillion

13
Are they guilty?

• Bands match, then that suspect is included in the
  group of individuals from whom the DNA evidence
  can come from
• Bands dont match, then suspect is excluded from
  individuals who could have contributed to the DNA
  evidence
• Inconclusive DNA possibly old or
  contaminated
• Can make no statement of guilt or influence, that
  is for a jury to decide

14
The Court Room

• When determining the admissibility of DNA
  evidence, courts consider the following
  questions
• The way the DNA sample(s) were obtained, gathered
  and preserved.
• The qualifications of the experts.
• The status of the laboratory.
• The reliability of the testing procedure.
• The majority of the time, the possibility of
  laboratory error is substantially larger than the
  possibility of a coincidental match. This is not
  because DNA laboratory work is particularly
  sloppy or unreliable. Instead, it is because the
  chance of a coincidental match is usually small.
  

15
Fake DNA Evidence

• Dr. Schneeberger, a Canadian doctor who hading
  been acquitted of sexually assaulting in 1994,
  had giving blood on three occasions. He had
  surgically inserted a Penrose drain, filled with
  foreign blood and anticoagulants in his arm.
• This only came to light when the police obtain a
  sample of his hair and Dr. Schneeberger was
  convicted in 1995.

16
Laboratories in Crisis

• Houston Police Departments Crime Lab
• 280 boxes contained evidence from 8,000 cases,
  items such as a human fetus, assorted body parts
  and a host of evidence had been stuffed into the
  boxes.
• This November, it was reviled that DNA evidence
  from 29 cases was either missing or had been
  destroyed.
• Retesting has put into question the conviction of
  15 cases including 4 death sentences.

17
Laboratories in Crisis cont.

• Cellmark
• Cellmark was awarded a 2.7 million three-year
  contract to provide DNA testing service to the LA
  Police Department, in January.
• In November, Sarah Blair a DNA analyst was fired
  for substituting data in control samples causing
  Cellmark to reanalyzed the evidence of several
  high-profile cases O.J. Simpson, JonBenet
  Ramsey, the Unabomber, etc.
• Other Labs
• West Virginia Fred Zain, the former director
  was facing fraud charges stemming from testimony
  provided, when he died in 2003.
• Marysville, Washington - In November 2001,
  Michael Hoover a forensic scientist at the State
  Patrol crime lab was sentenced to 11 months,
  after admitting to taking heroin from case
  evidence to ease him back pain.

18
Case Law

• The admissibility of scientific evidence in the
  federal court was established Frye v. United
  States, 1923. The Frye court put forward the
  "general acceptance" test.
• State v. Stills the Supreme Court of New Mexico
  approved the admissibility of PCR analysis
  stating that PCR analysis "has received
  overwhelming acceptance in the scientific
  community and the courts".

19
Areas of Statistical Calculations to Considered
in DNA Testing

• Population Genetics
• In calculating a probability labs employ
  databases, which are divided into broad racial
  categories comprising of the DNA profile of a
  few hundred individuals.
• These databases are built on the science of
  Population Genetics.
• Murder case in Vermont the defense argued that
  any DNA evidence should be inadmissible as there
  was no suitable reference database. The judge
  agreed, stating it is unclear which if any of
  the FBI database is appropriate for calculate
  probability of a coincidental match.

20
Areas of Statistical Calculations to Considered
in DNA Testing

• The Product Rule
• The product rule is employed in determining a
  match.
• Guidelines for the application of the product
  rule are detailed in the Recommendations 4.1 in
  the Second National Research Council Report (NRC
  II).
• The product rule requires an assumption of
  within (Hardy-Weinberg or HW) and between
  (linkage equilibrium or LE) locus independence,
  which cannot be exactly true.

21
The Ceiling Principle and the Modified Ceiling
Principle

• The National Academy of Sciences first
  recommended the use of the Ceiling Principle back
  in the late 1980s. The idea was to provide the
  most conservative estimate of probability but in
  the mid 1990 it fell out of favor
• Advances in DNA fingerprinting
• In the eighties the number of VNTRs used was
  small, four or less compared to today when the
  norm is to employ at least six loci.
• A Shift in the scientific landscape,
• Randjit Chakraborty and Kenneth Kidd defended DNA
  statistical analysis in their paper entitled The
  Utility of DNA Typing in Forensic Work. They
  asserted that even a small amount of migration
  (gene) across populations is quickly homogenized
  and contested Lewontin and Hartles ideas.
• In 1994, Bruce Budowle and Eric Lander published
  a paper in they concluded that any controversy
  had been resolved by the Technical Working Group
  on DNA Analysis Methods (TWGDAM) sponsored by
  the FBI and the Department of Justice.

22
Who was Hardy-Weinberg?
23
Cells, Chromosomes, Genes
23 pairs of chromosomes
Each set passed from parents, one from each
Inherited characteristics controlled by genes
24
Inheritance

• Genes can come in alternative forms, alleles

Each controls specific trait, example eye
color brown, green, blue
One allele from each parent
Dominant vs. Recessive
25
Recessive Sickle Cell Anemia
Allele causes shape of red blood cells to be
altered
2 copies - disease is manifested
1 copy - carrier
26
Dominant Huntingtons Disease
Progressive, usually mid 30s Involuntary
muscular action and dementia
Only 1 copy of allele needed
27
Two different Genes
What happens when at a given locus Both
Dominant Recessive alleles present
28
The Hardy-Weinberg Equilibrium
1908 Godfrey Harold Hardy - mathematician Wilhel
m Weinberg - physician
Mathematical equivalent of Mendels
observations on heredity
29
Phenotypes
For a gene with 2 alleles, A and a
p the population frequency of A
q the population frequency of a
30
Cross Product
A a A AA Aa a Aa aa
AA 25 Aa 50 aa 25
31
Genotypes
AA
Aa
Aa
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Restrictions
Population size infinite
Random mating
No selection for or against genotypes
No mutations
No net migration into or out of population
33
Applications of Hardy-Weinberg
DNA Forensics
Population Genetics
34
Forensics
DNA exclusion no match
DNA inclusion match but need to relate
to population frequencies
35
Frequency
Each allele has an observed frequency Use the
product rule
Blond hair 1/10 Blue eyes 1/10 Fair
skin 1/10 (.10) (.10) (.10) 1/1000
What about in Scandinavia? Issue of population
36
Confidence Levels
95 confidence limit, true value has only 5
chance exceeding upper bound
If pattern 1/100 Frequency .01 Upper
confidence limit 4.7
37
Population differences
A a Group I - 90 .5 .5 Group II -
10 .9 .1
with random mating AA Aa aa Group
I .25 .50 .25 Group II .81 .18 .01
38
The Results, not that different
Observed genotype AA(.9)(.25)(.1)(.81) .306 A
a(.9)(.5) (.1)(.18) .468 aa(.9)(.25)
(.1)(.01) .226
Without regard to population substructure A(.9)(
.5)(.1)(.9) .54 a(.9)(.5)(.1)(.1) .46
Hardy-Weinberg proportions AA(.54)(.54) .2916
Aa2(.54)(.46) .4968 aa(.46)(.46) .2116
http//www.dartmouth.edu/chance/teaching_aids/boo
ks_articles/DNAtyping/node1.html
39
Population Genetics
Sickle Cell Anemia observed frequency 0.09
p q 1
q population with SCA .09 p
population with out SCA .91
40
Expected Frequencies
Genotype frequencies
41
Conclusion
Hardy-Weinberg Equilibrium is a powerful tool
Pay attention to its constraints
42
References

• i http//www.bbc.co.uk/crime/caseclosed/coli
  npitchfork.shtml
• ii http//en.wikipedia.org
• iii On Conveying the Probative Value of DNA
  Evidence Frequencies, Likelihood Ratios, and
  Error Rates, by Jonathan J. Koehler, The
  University of Texas at Austin, 20th March 1996
• iv http//www.medterms.com/
• v http//www.cnn.com/2004/TECH/science/11/18/
  dnalab.ap/index.html
• vi http//www.law-forensic.com/cfr_gen_art_24.
  htm
• vii http//www.cellmark-labs.com/
• viii Labs Placed Under a Microscope, by Robert
  Tanner, The Washington Post, July 27, 2003
• ix http//www.illinoisbar.org/Member/mar00lj/p
  134.htm2
• x http//www.kscourts.org/ca10/cases/2001/03/
  00-2475.htm
• xi http//www.forensic-evidence.com/site/EVID/
  EL_dna_instr_bad.html
• xii http//www.fathom.com/course/21701758/sessi
  on4.html
• xiii Interpreting DNA Mixtures Based on the
  NRC-II Recommendation 4.1, by Wing K. Fung and
  Yue-Qing Hu, Forensic Science Communication,
  Volume 2 - Number 4, October 2000
• xiv Commonwealth v. Lanigan, 419 Mass. 15
  (1994)
• xv Utility of DNA Typing in Forensic Work, by
  Randjit Chakraborty and Kenneth Kidd, Science
  20th December 1991, Vol, 254, Issue 5039
• xvi http//www.nas.edu/
• xvii Use of DNA Profiles in Criminal
  Proceeding, Alaska Judicial Council, December
  1996
• xviii Dr. Shockley and Mr. Hyde, by Joseph
  Galloway et al, News World Report, 28th Aug
  1989
• xvii http//www.law.berkeley.edu/journals/btlj/
  articles/vol8/Denemark/html/reader.html