DNA profiling

What is it?

• What do you know about DNA Profiling?– How does it work?– From what sources can DNA for profiling be

obtained? – How reliable is it?– How is enough DNA obtained when little is available

from the crime scene?– What, exactly, is a DNA profile or fingerprint?

Origin of forensic techniques

• Examination of biological evidence– Karyotyping – analysis of chromosomes – Blood typing – excludes suspects, determines if

blood at crime scene came from one or multiple suspects

• Most techniques originated for medical diagnosis or treatment

Karyotyping

DNA fingerprinting

• DNA profiling – used in criminal and legal cases to determine identity or parentage– DNA extracted from very small amounts of

biological evidence• Drop of blood• Single hair follicle

– Can accurately predict whether an individual can be linked to a crime scene or excluded as a suspect

Function and Structure of DNA

• DNA – Deoxyribonucleic acid• Contains the genetic material of the cell• Instructions to make proteins and to replicate• Chromosome – structures made up of

molecules of DNA

DNA molecule

• Double-helix structure – composed of two strands

• Sides of the ladder – backbone of sugar and phosphate molecules

• Rungs of the ladder – nitrogenous bases

The Different DNA Bases

• Adenine (A), Cytosine (C), Guanine (G), Thymine (T)

• Binding– A with T– C with G

• Complimentary Strands:– Strand: CGTCTA– Compliment: GCAGAT

Where does your DNA come from?• 23 pairs (46)of chromosomes in human cells– Exception is human reproductive cell

• Nuclear DNA – in nucleus– One chromosome from mother, one chromosome

from father– Virtually identical in all cells of the body

• Mitochondrial DNA – in cytoplasm– Exists in a circular loop– Passed on only from mother

Genes and Alleles

• Genes – DNA sequences that determine our inherited characteristics or traits

• Allele – one of two or more alternative forms of a gene– One comes from mother, one from father– Ex. One allele codes for normal hemoglobin– One allele codes for abnormal hemoglobin

Human Genome Project

• 13 year project to determine the order of bases on all 23 pairs of human chromosomes

• Helps to know where on a specific chromosome DNA codes for production of a specific protein

• Can help to treat and/or cure genetic diseases and disorders

Human Genome Project

• 3 million base pairs in human DNA• Exons (encoded DNA) – contains DNA used to

make proteins or other molecules • Introns (unencoded DNA, “junk DNA”) – isn’t

involved in protein making, but important for gene splicing– More than 98.5% of genome is junk DNA– Mutations in the non-coding are tolerated and can

accumulate with no effect on the organism

DNA Identification

• Polymorphisms – unique patterns of repeated base sequences that identify individuals– Found in introns (junk DNA)

• Different repeated sequences appear in different places in the genome of each individual

• Two types of repeats in noncoding sections:– VNTRs– STRs

Discovery of DNA profiling

• 1984 – Alec Jeffreys: University of Leicester• Developed technique for DNA fingerprinting– Appears as a pattern of bands on x-ray film– Each individual has a unique DNA band pattern

• Shows:– Same individual– Related individuals– Unrelated individuals

Alec Jeffreys

VNTR - Variable Number of Tandem Repeats

• Short sequences of DNA that are repeated multiple times– Number of copies of the same repeated base

sequence varies among individuals– CATACAGAC• 3 copies in one individual, 7 in another

– Length of VNTR can be from 9 to 80 bases in length

STR – Short Tandem Repeats• Short sequence of DNA with a high degree of

polymorphism– Usually 2 to 5 base pairs in length– GATA

• Ex- GATAGATAGATAGATA

– Polymorphisms result from different # of copies of repeat element

– Becoming preferred method of analysis• Accurate• Small and partially degraded samples may be used• STRs much shorter than VNTRs so much easier to separate

DNA Profile

• Multiple VNTRs and STRs examined • Purpose:– Tissue Matching – connecting a suspect to a crime

scene– Inheritance Matching – each band in a child’s DNA

fingerprint is present in at least one parent

Population and Genetics

• Population genetics – the study of variation in genes among a group of individuals

• Population studies conducted to calculate the percentage of people who have a particular allele.– The greater the number of people, the more

accurate a statistics will be– Ex –probability of blood type of a suspect matches

crime-scene evidence, must refer to a database with info from a representative population.

Sources of DNA at Crime Scenes

• Blood (White blood cells)• Semen (Sperm cells)• Hair with roots (Hair follicle cells)• Skin, dandruff (Skin cells)• Sweat stains (Skin cells sloughed off)• Vaginal fluids (Mucosal surfaces)• Nasal secretions (Mucosal surfaces)• Urine (Mucosal surfaces)• Feces (Digestive system cells)• All individual evidence!!!

PCR – Polymerase Chain Reaction

• Trace evidence – when the amount of evidence left at the crime scene is very small

• Problem – evidence may be totally consumed during testing

• Solution: PCR - generates multiple identical copies from trace amounts of original DNA evidence

Collection and Preservation of DNA Evidence

• Contamination issues – when DNA from another source is mixed with DNA relevant to a case– Ex – coughing, sneezing, touching mouth, nose,

eyes • Keep evidence dry and cool• Moisture, sunlight can damage and denature

DNA

Precautions1. Wear disposable gloves and change them often.2. Use disposable instruments for handling each

sample.3. Avoid touching the area you believe DNA may exist.4. Avoid talking, sneezing, and coughing over

evidence.5. Avoid touching your face, nose, and mouth when

collecting and packaging evidence.6. Air-dry evidence thoroughly before packaging.7. Put evidence into new paper bags or envelopes.8. If wet evidence cannot be dried, it may be frozen.

Steps of DNA Fingerprinting

1. Extraction 2. Restriction fragments3. Amplification4. Electrophoresis

1. Extraction

• DNA is extracted from cells• Cells are isolated from tissue• Cell membrane and nuclear membrane are

broken down– Let’s watch:

2. Restriction Fragments

• Fragments of varying lengths formed when DNA is cut

• Restriction enzymes – recognize a unique pattern of DNA bases and will cut DNA at specific location– “molecular scissors”– Often used and produced by bacteria (to destroy

nucleic acid of enemies)– Many different restriction enzymes , each binds to

specific recognition site

Restriction Enzymes

Restriction enzyme Hind III• AACGTT• Hind III cuts

between the two AA bases

• some of the fragments produce VNTRs

Recombinant DNA

• DNA that has been artificially created• Cultured cells (E. coli, yeast, mammalian cells)

transformed with a human gene manufacture products for human therapy:– Insulin– Human growth hormone– Parathyroid hormone– Factor VII, Factor IX for hemophelia

3. Amplification

• Making more copies of the DNA • This is done for several reasons:– Most evidence samples from a crime scene

contain very little DNA– It is a standard or control (i.e. important for

Daubert challenges) – one needs to argue that the same amount of DNA is used in each lab, by each lab technician and every time sample is processed

Amplification

• PCR – Polymerase Chain Reaction• Amplify certain pieces of DNA that contain

VNTRs and STRs• PCR Machines, or thermocyclers:– use repeated cycles of heat and cooling to

replicate the DNA – use many of the same enzymes found in cells

which facilitate DNA replication

4. Electrophoresis• Method of separating the molecules within an electric field

based on size of DNA fragments• Negatively charged DNA moves through gel with a current• Fluorescent dye is binded to DNA • Smaller fragments move faster than larger fragments

Electrophoresis• Steps:

1. Micropipette used to place DNA into separate chambers or wells across top of gel• Control – DNA Ladder or Standard DNA

2. Electric current passed through gel3. Process complete when DNA fragments have

migrated through gel4. Visualize DNA fragments through probes

DNA and Probes

• Radioactive probes – used to view specific DNA sequences in a DNA fingerprint

• STR analysis – only fragments that were amplified will be on gel– Fluorescent stain in the liquid the gel is bathed in– UV light source used to cause DNA to fluoresce

and make bands visible

DNA and Probes• DNA probe – made up of a synthetic sequence of

DNA bases that is complimentary to a small portion of DNA strand– AAGCTTA, radioactive probe – TTCGAAT

• Next step is to view DNA fingerprint• Depends on the chemical used to label the probe– Fluorescent dye – viewed under UV light– Radioactive isotope – X-ray film produces

autoradiograph• Technique called Southern Blotting

Paternity

• DNA fingerprint is a combination of DNA from mother and father– Position and width of bands is significant– All bands have to match exactly

• Greater the number of probes used, the greater the accuracy– Six to eight probes common for DNA fingerprint

Analysis of DNA Fingerprints

• Combined DNA Index System (CODIS) – electronic database of DNA profiles– Uses 13 loci– Tetrameric repeats

• States maintain DNA index of individuals convicted of rape, murder, child abuse

• DNA profiles from a crime scene can be entered into CODIS to identify possible suspects

A Sample Profile

• By combining the frequency information for all 13 CODIS loci, the frequency of this profile would be 1 in 7.7 quadrillion Caucasians

As the technology gets smarter, so too do the criminals

• A physician in Canada eludes authorities for years• Accused of drugging and sexually assaulting patients, DNA

profiles from semen samples from the assaulted women do not match Dr. Schneeberger

• Blood was drawn on 3 occasions in 1992, 1993 and 1996, but never came back as a match

• Finally police obtain blood from a finger prick, swabbed the inside of his cheek and took hair samples

• The results matched the DNA from the semen of the victims• How did he get away with it?

As the technology gets smarter, so too do the criminals

• On the previous 3 occasions, blood was drawn from the same arm

• The last time the blood was drawn, the technician stated that the blood looked brown and “old”

• Schneeberger had surgically implanted a piece of rubber tubing in his arm and filled it with stored blood from a patient

Its Uses

• Identification of remains

The Angel of Death: Josef Menegle

• Josef Mengele was a Nazi war criminal notorious for grotesque human experiments that he carried out at the Auschwitz concentration camp.

• After the Second World War he fled from the Allies and escaped to South America. The fugitive succeeded in living out the rest of his days without being caught.

• In 1985 investigators went to the cemetery of Nossa Senhora do Rosario in the small Brazilian town of Embu to dig up the skeleton of a man who had been drowned in a swimming accident six years previously.

• Using DNA extracted from blood provided by Mengele’s wife and son, it was concluded that it was more than 99.94% certain that the skeleton was Mengele’s.

Paternity Cases

• Who’s your daddy?

1.

2.

1. 2.

Homicide or Rapes:

OJ Simpson

Exoneration

• Kirk Bloodsworth– Convicted in 1985 for the

rape and strangulation of a 9-year old girl and sent to death row

– In 1992, defense attorneys were successful in having a dime-sized semen stain on the girl’s underpants tested against Bloodsworth’s DNA

– He was exonerated

Exoneration

Uses of DNA fingerprinting

• Crime-scene DNA matching with suspect’s DNA

• Paternity and maternity determination• Identify family members and relatives• Suspect elimination• Exoneration of falsely accused and imprisoned• Identification of human remains