Forensic Biology

Rachel NeagleVirginia Department of Forensic Science

© 2014 Virginia Department of Forensic Science

Outline• Introduction to DNA• Forensic Biology Lab Processing

– Body Fluid Identification– Nuclear DNA Testing

• Analysis– Statistics

• Data Bank• Additional DNA Testing

Introduction to DNA

What is DNA?• Deoxyribonucleic Acid

– Chemical substance found in our bodies that is responsible for determining all aspects of our physical make-up

– “Blueprint of Life”

What is DNA?• Deoxyribonucleic Acid

– The molecular basis of inheritance• ½ from mother and ½ from father

– >99% of DNA is shared between individuals

– Identical siblings share the same DNA

Identical DNA??

Where is DNA found? Human Cell

Mitochondria• powerhouse of

cell

• Mitochondrial DNA

• high copy number

Nucleus• brain of cell

• Nuclear DNA

• high power of discrimination

www.cstl.nist.gov/bioteh/strbase

Where is DNA found?• Blood (WBCs)• Semen• Saliva• Perspiration/Skin cells• Tissue• Hair• Bone• Teeth• Urine

Structure of DNA

• Chromosomes– Tightly wound,

supercoiled DNA

• 23 pairs of chromosomesh

ttp://www.ncbi.nlm.nih.gov/genome/guide/

Forensic Biology Lab Processing

Forensic Biology Lab Processing

• Consists of both serological examination and DNA analysis

• Examines evidence for:– The presence/absence of body fluids/tissue (blood, semen,

urine, perspiration, etc)– Determine DNA profile

Virginia DFS Western Lab

Virginia DFS Western Lab

Biological Evidence

• Body fluids &/or tissue– Blood– Semen– Saliva– Hairs– Bone/teeth– Skin/tissue– Sweat– Urine– Feces

Forensic Biology Evidence

Examples of Touch Evidence

• Keys• Gun grips & triggers• Door & drawer

handles• Light switches• Countertops• Gear shift knobs• Steering wheels• Cigarette lighters

Blood Testing• Chemical Testing

– Phenolphthalin (Kastle-Meyer)– Tetramethylbenzidine– Combined Phenolphthalin Tetramethylbenzidine (PTMB)– Leucomalachite Green (LMG)

• Chemiluminescent Testing– Luminol– Bluestar®

• Immunological Testing– Hematrace®

• Microscopic Exam– Teichman– Takayama

Blood Testing• Chemiluminescent Testing

– Luminol– Bluestar®

• Chemical Testing – Combined Phenolphthalein Tetramethylbenzidine

Phenolphthalein Tetramethylbenzidine

• Alternate Light Source

Semen Testing

Semen Testing

• Chemical Testing– Acid Phosphatase

(AP)– Presumptive Test

Semen Testing

• Microscopic examination for spermatozoa

Semen Testing• P30 (prostate specific antigen)

2nd pink line in the Test Area indicates a positive result

Urine, Feces and Saliva Testing• Urine

– Urease Test –breakdown of urea by urease in presence of heat causes basic reaction

• Feces– Edelman’s Test – tests for the presence of urobilin

which is a breakdown product of bilirubin• Saliva

– Phadebas Test- tests for the presence of amylase

These tests require a significant amount of sample

DNA Testing

• DNA Testing– Extraction and purification– DNA quantitation– PCR set-up

• Analysis of results

DNA Extraction• DNA is removed from the

cellular material and purified• Chemicals + Heat

DNA Extraction

• Mixed stains– Sperm cells +

epithelial cells– Differential

Extraction

DNA Extraction

Quantitation

• Estimates how much DNA is present in each sample

• Information used for processing

PCR

• Polymerase Chain Reaction

• Makes millions of copies of a specific region(s) of DNA (AKA amplification)

• Similar to Xeroxing

PowerPlex® 16

15 STR loci + Amelogenin (sex determination)

Automation of DNA Testing

• Advantages:– Increase sample

production/ throughput

– Increase time for examiner to conduct other duties (screen additional cases)

– Decrease risk of contamination

– Increase consistency of results

3100xl Instrument for DNA Analysis

• DNA sample injected automatically onto a capillary column

• DNA separated by electrophoresis

• Data automatically collected

DNA Analysis• Separation of the copied regions based on

size to determine differences and determine a profile of the contributor(s)– Pieces with more repeats, larger, move

slower through gel-like substance– Represented by peaks on a graph

(electropherogram)

• The DNA profile

Nuclear DNA Analysis

STRs – Short Tandem Repeats• Non-coding regions of DNA• Alleles assigned numbers based on the number

of 2-7 bp repeated segments of DNA present• Examine 15 different STR loci + gender marker

Nuclear DNA Analysis

AGAT AGATAGAT

AGAT

AGAT

AGAT

AGAT

AGAT

AGAT

AGAT

6

4

DNA Profile = 4,6

One piece inherited from mom, the other from dad

Nuclear DNA Analysis

• Mixture Profiles– DNA profile resulting from more than one

individual contributing to a sample– Three or more peaks (alleles) generally

being detected at multiple areas– Major/minor contributors

Conclusions

• Included (Not eliminated)– Individual cannot be eliminated as a contributor

of a DNA profile or to a DNA mixture profile• Eliminated

– Individual is not the source of the DNA detected• No Conclusions

– Cannot make a determination relative to individual in question

Statistics

• Provides a measure of the weight to the evidence– How common or uncommon is a particular

profile in the population?– How often you would expect the evidence

profile to be found in the population?• (based on allelic frequencies)

Statistics

• Calculated when an individual is included as a possible contributor of the evidentiary DNA profile

• Numerical probability of approximately how rare or how common a given DNA profile is in the population

Factors which limit effectiveness of DNA evidence

• Contamination– Direct contact with another source of DNA– Can occur before, during, or after evidence

collection• Degradation

– High temperatures, humidity– Bacterial growth, natural decaying process

Factors which limit effectiveness of DNA evidence

• Probative value– Evidence left by someone not related to the crime– Evidence obviously left by someone related to

crime• Availability of reference samples

– Mixtures without reference(s) have little value– Complex mixtures without reference(s) typically

have no value

DNA Data Bank

The collection and DNA analysis of blood and buccal samples from convicted felons and Arrestees

DNA profiles obtained from evidence samples are used for comparison to unsolved cases and samples from convicted offenders and arrestees

DNA Data Bank

CODIS

• Software owned and operated by Federal Bureau of Investigation

• Comprised of samples from the federal databank and samples from all 50 states

• Samples include:– Offender– Arrestee– Forensic– Missing persons

NDIS Statistics

• As of June, 2014– Over 11,015,147 offender samples– Over 1,922,415 arrestee samples– Over 565,159 forensic samples– Produced over 250,809 hits

For statistics:http://www.fbi.gov/about-us/lab/biometric-analysis/codis/ndis-statistics

Additional DNA Testing

• Y-STR analysis

• Mitochondrial Analysis

Y-STR Analysis

• Similar to nuclear DNA analysis except focuses on the Y-chromosome

• Paternally inherited• Commercially available amplification kit

for 17 areas on the Y-chromosome

Y-STR Analysis

• Advantages– Extreme female-male ratio cases

• With or without sperm presence– Male-male mixture cases

• Disadvantages– Standards needed for comparison– Not unique to individual-paternally inherited

Where is DNA found? Human Cell

Mitochondria• powerhouse of

cell

• Mitochondrial DNA

• high copy number

Nucleus• brain of cell

• Nuclear DNA

• high power of discrimination

www.cstl.nist.gov/bioteh/strbase

Mitochondrial DNA Analysis

• Mitochondrial DNA– Mitochondria: Cytoplasm– >1000 copies per cell– Maternally inherited

National Institutes of Health Genome.gov

Mitochondrial DNA • Advantages

– Mulitiple copies per cell- >1000 copies– Reference samples more easily obtained

• Maternal relatives– Harder to degrade than nuclear DNA– Found in lower quantity or highly degraded samples

• Hair shafts• Bone

– Body ID, missing persons and mass casualty cases

Photo courtesy of Robert Quick, Norfolk PD

http://commons/wikimedia.org/wiki/Image:CID_post-impact_1.jpg

Mitochondrial DNA

• Disadvantages– Testing is labor intensive and costly– Maternally inherited

• Not unique to an individual• Statistics generally 1 in 100’s to 1 in 1000’s

– Reference standards required because most samples cannot be searched in database

– Not applicable to mixture samples

Acknowledgements

• Virginia DFS co-workers• www.dfs.virginia.gov• www.fbi.gov• www.nih.gov• www.nist.gov

Questions?