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Forensic DNA Analysis

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Title: Forensic DNA Analysis


1
Forensic DNA Analysis
  • For today, 11/5 Lecture
  • Pass back papers
  • Keep your graphic organizer on your desk
  • If you did not turn yours in yesterday, turn in _at_
    the end of the period.
  • Put your gel electrophoresis paper in your binder
    we will look at that tomorrow.
  • Fill in notes on your graphic organizer where you
    can, otherwise add them in to you class notes

2
  • Objectives of Forensic DNA Testing
  • To link an individual to a crime scene/criminal
    act
  • To exonerate suspects
  • To identify victims of mass disasters
  • http//www.innocenceproject.org/
  • Site with interesting lessons on DNA analysis
    http//www.nfstc.org/pdi/Subject00/pdi_s00.htm

3
Terms to know
  • in vitro
  • Refers to a life process occurring outside of a
    living organism or cell, typically in a test tube
    or petri dish
  • PCR (polymerase chain reaction)
  • An in vitro method for replicating/copying DNA
  • Thermocycler
  • A piece of equipment used in PCR to heat and cool
    the samples

4
Terms to know contd
  • Restriction enzyme
  • An enzyme that cuts DNA at or near a specific
    sequence of nucleotides
  • Power of discrimination/exclusion
  • Power of a test to eliminate a certain of the
    population from being biologically related to an
    individual
  • If a paternity test using blood typing has a
    power of exclusion of 30, only 30 of the male
    population could be eliminated from being the
    father of said child
  • Loci
  • Specific location of a gene or DNA sequence on a
    chromosome

5
Early 1980s- mid 1990s Restriction Fragment
Length Polymorphism (RFLP)
  • DNA sample is exposed to restriction enzymes that
    cut at specific sequences around VNTRs
  • Fragments are different lengths due to the of
    repeats
  • These fragments are sorted using DNA
    electrophoresis
  • http//www.pbs.org/wgbh/nova/education/body/create
    -dna-fingerprint.html
  • Power of discrimination in the range of 106-108
    for a six probe analysis

Sir Alec Jeffreys
6

Mechanisms for RFLPs
7
Mid-1980s The Colin Pitchfork Case
  • Two young women raped and murdered in Narborough,
    England
  • 5,000 local men are asked to provide blood/saliva
    samples
  • 1st exoneration and conviction on forensic DNA
    evidence

8
RFLPs contd
  • Benefits
  • Drawbacks
  • High power of exclusion (gt or 99.99)
  • Reaches high power of exclusion while only
    needing to analyze a few (6) loci
  • Requires large samples of blood
  • Long strands of bases (1000bp) and tend to be
    broken or damaged
  • Can take up to 8 weeks for analysis
  • DNA evidence at a crime scene is typically of a
    small quantity and degraded

9
PCR To The Rescue!
  • Enabled DNA replication in vitro so sample
    sizes from a crime scene could be amplified
  • First forensic application DQa locus,
  • Advanced applications are multi-plexed with
    Polymarker loci using dot-blot detection method
  • Works with lower quantity (1-2ng), lower quality
    samples
  • Power of discrimination goes from 102-106...not
    good enough for databasing

Dr. Kary Mullis
10
The Current Method of Choice STRs (short tandem
repeats)
  • Non-coding, tetranucleotide sequences (3-7 bases
    long) which vary greatly from person to person in
    the number of repeating units
  • Requires lt1ng of DNA to type 13-15 STR loci
  • The of repeats of STRs at 13 different,
    distinct locations on the chromosomes
  • Human Genome Project uses STRs

11
The Process In a Nutshell
  • Amplified DNA samples are injected into a
    capillary
  • Fluorescent tags on the DNA fragments are excited
    by a laser
  • As they pass a window in the capillary, the
    fluorescence is recorded by a camera, and this
    signal is converted into a peak by the computer
    software.
  • http//www.dnalc.org/view/15983-Today-s-DNA-profil
    e.html

12
STRs contd
  • Benefits
  • Drawbacks
  • Small and easily amplified through PCR
  • Can detect biological relationships to both
    mother and father
  • High power of discrimination
  • 1014-1023 (world population is 109)
  • Creation of CODIS
  • See p. 328 in text, Table 9.1 for STRs used in
    CODIS
  • Newer, still refining

13
The Combined DNA Index System (CoDIS)
  • A database of DNA profiles from violent felons
    and crime scene samples
  • Laws concerning who is eligible for the database
    vary from state to state
  • Database currently contains about 2,038,470
    felons and 93,956 crime scene profiles (19,00
    hits so far)

14
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15
The Dark Side of CODIS(What the FBI doesnt want
you to know.)
  • DNA mixtures and degraded DNA profiles have lead
    to spurious matches
  • Stringent laws explicitly prohibit databasing
    innocent people
  • Adding arrestees to database violates presumption
    of innocence
  • However, the prosecution rate on case to offender
    matches is shockingly low! (10)

16
LAPD CoDIS Statistics
  • 177/142 Case-to-Offender matches
  • 100 Case-to-Case hits with 42 as yet unidentified
    suspects
  • 28 DA rejects
  • 9 Convictions charges filed in 28 more 4
    defendants plead guilty

17
Specialized PCR-based systems
  • mtDNA
  • Y-STRs
  • SNPs

18
Mitochondrial DNA (mtDNA)

19
mtDNA contd
  • Benefits
  • Drawbacks
  • Single-cell sensitivity because each cell
    contains 1000 mitochondria
  • Especially useful for shed hairs, burnt remains
  • Can be used to establish kinship directly because
    entire complement of mtDNA is maternally
    inherited
  • Single-cell sensitivity because each cell
    contains 1000 mitochondria very high
    contamination risk!
  • Lower power of discrimination - maternal
    relatives all share the same mtDNA

20
Y-STRs
  • Problem
  • 99 of violent crimes are committed by men
  • DNA Mixtures of male suspect and female victim
    can pose an analytical challenge, especially when
    the female contribution is much greater than the
    male preferential amplification
  • Solution
  • Test for markers found only on the Y-chromosome.
    Only male DNA is amplified!

21
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22
Y-STRs
  • Khan Argument
  • Lower power of discrimination
  • Paternal relatives all share the same Y-STR
    haplotype (Wicked Uncle Ernie Defense)
  • 10 of Central Asian males share the same Y-STR
    haplotype, thought to belong to Genghis Khan

23
SNPs (Single Nucleotide Polymorphisms)
  • Point mutations (base substitutions) found in 1
    or more of the population
  • 1.8 million identified in human genome
  • Detected on micro-array plates with fluorescent
    tags (all or nothing response)
  • 50 SNPs provides same power of discrimination as
    13 STR loci
  • Certain SNPs used as predictors of
    ancestry/ethnicity by a private sector lab (DNA
    Witness)

24

SNP Flaws
  • Privacy issues - unlike STRs, SNPs can be
    correlated with susceptibility/resistance to
    diseases
  • Requires a relatively large quantity of DNA for
    robust assay
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