Comprehensive Study Guide for DNA Forensics and Profiling

Foundational Perspectives and Structural Chemistry of DNA

  • Lewis Thomas Perspective: The physician and author Lewis Thomas noted that the capacity of DNA to "blunder slightly" (mutate) is a marvel of biology, as it allowed for evolution beyond anaerobic bacteria to the complexity of human life and culture, such as music.

  • Historical Pioneers: The understanding of DNA as a double-stranded molecule is attributed to the work of James Watson and Francis Crick, along with the critical contributions of Dr. Rosalind Franklin through X-ray crystallography.

  • DNA as a Polymer: DNA is a long-chain polymer found in the nucleated cells of living organisms. It functions as a repository for genetic information.

  • Molecular Structure:

    • Double Helix: The structure consists of two coiled DNA strands.

    • Nucleotide Composition: The basic units of DNA are nucleotides, which consist of three components:

      • A sugar molecule (specifically deoxyribose).

      • A phosphate group.

      • A nitrogen-containing base.

    • Nitrogenous Base Pairing: There are four bases, which follow strict complementary pairing rules:

      • Adenine (AA) always pairs with Thymine (TT).

      • Guanine (GG) always pairs with Cytosine (CC).

    • Human Uniformity: In humans, the order of these base pairs is approximately 99.9%99.9\% identical across all individuals.

Forensic Utility and Evidence Applications

  • Individual Evidence: Because every person’s DNA is unique (with the exception of identical twins), it serves as a form of individual evidence.

  • Crime Scene Utility:

    • Suspect Linkage: DNA can link a suspect to a crime scene or eliminate them, functioning similarly to a fingerprint.

    • Victim Identification: DNA from relatives can be used to identify deceased victims.

    • Serial Crime Linking: It can connect different crime scenes by identifying the same perpetrator across local, state, and national jurisdictions.

  • Case Refutation and Alibis:

    • DNA can place individuals in specific rooms or homes where they claimed not to have been.

    • It can refute claims of self-defense by identifying who held a specific weapon.

    • It can shift a legal defense from a total alibi to a claim of consent.

Biological Sources of DNA

  • Cellular Origins: DNA is found in all nucleated body cells.

    • White Blood Cells (WBCs): Provide the DNA in blood samples; Red Blood Cells (RBCs) lack nuclei and therefore do not contain nuclear DNA.

    • Body Fluids and Tissues: Semen, saliva, urine, teeth, bone, and soft tissue are all viable sources.

    • Buccal Cells: Cells from the inner cheek are the most abundant source for reference samples.

    • Hair: DNA is found specifically in the hair roots.

Comparative Analysis: Nuclear vs. Mitochondrial DNA

  • Nuclear DNA (nDNA):

    • Location: Found within the nucleus of the cell.

    • Inheritance: Inherited from both the mother and the father.

    • Quantity: Each cell typically contains only one nucleus.

    • Structure: Organized into 2323 pairs of chromosomes; contains approximately 3×1093 \times 10^{9} bases and over 20,00020,000 genes.

    • Reproductive Context: nDNA is located in the head of the sperm. During conception, the head enters the egg to unite with the nucleus.

  • Mitochondrial DNA (mtDNA):

    • Location: Found within the mitochondria in the cytoplasm.

    • Structure: Exists as a circular DNA molecule; contains approximately 16,50016,500 bases and 3737 genes.

    • Inheritance: Inherited strictly from the mother (maternal lineage).

    • Abundance: Each cell contains hundreds to thousands of mitochondria, meaning there are many copies of the mitochondrial genome per cell.

    • Stability: It is very hardy and can be recovered from old bones, teeth, and hair. It has a very low mutation rate (roughly once every 6,5006,500 years).

    • Reproductive Context: mtDNA is located in the tail of the sperm, which falls off during conception and is not passed to the offspring.

    • Forensic Rigor: mtDNA analysis is more time-consuming, rigorous, and costly than nDNA testing. It is typically used when nDNA is too degraded or unavailable.

Polymorphisms and DNA Profiling

  • Coding vs. Non-Coding Regions:

    • Encoded DNA: Only 3%3\% of human DNA codes for proteins; this portion is largely the same for everyone.

    • Non-Coding DNA: 97%97\% of the genome is non-coding and repetitive. These regions are called polymorphisms (meaning "many forms").

    • Variation: Most individual variation is found in non-coding regions where specific base sequences repeat. Only about 3×1063 \times 10^6 bases differ between individuals.

  • Purposes of DNA Fingerprinting (Profiling/Typing):

    • Tissue Match: Determining if two samples have identical banding patterns (originating from the same person).

    • Inheritance Match: Ensuring that every band in a child’s DNA fingerprint matches a band present in at least one parent.

  • General Forensic Uses:

    • Identifying potential suspects or exonerating the wrongly accused.

    • Identifying remains in mass disasters (e.g., WWII, 9/11).

    • Establishing paternity and relatedness.

    • Matching organ donors for medical procedures.

Methodology: The 4 Steps of DNA Fingerprinting

  1. Extraction: Removing the DNA from the cell.

    • The cell membrane is disrupted using a detergent.

    • Isopropyl alcohol is added to separate the DNA from other components; the DNA moves into the alcohol layer and is spooled onto a glass pipette.

  2. Restriction Fragments (RFLP): Using restriction enzymes, often called "genetic scissors," to cut DNA into fragments of different lengths.

  3. Amplification (PCR): Using Polymerase Chain Reaction to make millions of copies of a defined DNA segment from a minimal sample (e.g., a speck of blood).

  4. Electrophoresis: Separating DNA fragments by size using an electric current moving through a gel.

    • Molecules sort by size in a porous gel maze.

    • The largest DNA fragments move the slowest and shortest distance.

    • The smallest DNA fragments move the fastest and farthest distance.

    • Visualization: Radioactive probes are attached to the fragments to create an X-ray record (autoradiograph) of the bands.

Forensic Interpretation and Outcomes

  • Gel Electrophoresis Analysis: Interpretation involves comparing lanes (e.g., Crime Scene vs. Suspects vs. Controls/Standards).

  • Possible Results:

    • Match: The DNA profiles are identical. The lab then determines the statistical frequency of that profile in the general population.

    • Exclusion: Genotype differences indicate the samples definitely originated from different sources.

    • Inconclusive: The data is insufficient to support either a match or an exclusion.

Short Tandem Repeats (STR)

  • Definition: STRs are short sequences of 22 to 1010 base pairs that repeat multiple times in the non-coding regions of the genome.

  • Advantages: STR analysis is faster, requires smaller/more degraded samples than RFLP, and is highly exclusionary.

  • STR Procedure Example (TH01 Locus):

    • 1. Extract the gene TH01 (which has seven variants with an A-A-T-G repeat).

    • 2. Amplify via PCR.

    • 3. Separate via gel electrophoresis.

    • 4. Determine the number of repeats based on migration distance.

  • Visualization: STRs are graphed as peaks. Each peak represents a DNA fragment size. People have two STR types for each gene (one from each parent).

  • Probability: By testing multiple STR loci, the mathematical probability of two people sharing the exact profile becomes minuscule.

Comparison of DNA Technologies: RFLP vs. STR

  • RFLP (Restriction Fragment Length Polymorphism):

    • Uses restriction enzymes to cut DNA into variable-length fragments.

    • Variation is caused by individual genetic differences in the length of these fragments.

    • It is older, slower, and no longer stored in modern databases like CODIS.

  • STR (Short Tandem Repeats):

    • Involves repeating sequences in non-coding regions.

    • More efficient with small or degraded samples.

    • Pairs well with automated systems like CODIS.

National and International Databases

  • CODIS (Combined DNA Index System):

    • The FBI’s program for linking serial crimes and unsolved cases.

    • Launched in October 19981998.

    • Links all 5050 US states.

    • Marker Requirements: Originally required 1313 core STR loci (D8S1179, D21S11, D5S818, CSF1PO, D3S1358, TH01, D13S317, D16S539, TPOX, D18S51, vWA, D7S820, FGA). This was recently expanded to 2020 core loci (adding D1S1656, D2S441, D2S1338, D10S1248, D12S391, D19S433, and D22S1045).

  • International Standards: The 2008 EU legislation (Pr̈m Treaty) requires member states to share DNA data for cross-border crime investigation.

Legal Cases and Genetic Genealogy

  • Sir Alec Jeffreys: Credited with developing DNA profiling using RFLP in September 19841984.

    • Case of Richard Buckland (19851985): Jeffreys used DNA to prove that a 17−year−old17-year-old was not responsible for a second rape he had been accused of, despite a confession.

    • Case of Colin Pitchfork: DNA evidence was used to identify and convict Pitchfork for the crimes.

  • The Golden State Killer (Joseph James DeAngelo Jr.):

    • Arraigned April 2727, 20182018; suspected of 1212 homicides, 4646 rapes, and 120+120+ burglaries from 1974−19861974-1986.

    • Genetic Genealogy: Investigators used sites like Ysearch.org and GEDmatch to find distant relatives, built a family tree, and matched discarded DNA from a public place to crime scene evidence.

  • Ethics of Genealogy Sites: Unlike federal databases, which have strict limits on searches, genealogy sites can be searched frequently. Uploading a profile may surrender the privacy of extended family members.

Economic Evolution of DNA Testing

  • The Human Genome Project (1990−20031990-2003): Cost approximately $2.7 billion\$2.7\text{ billion}.

  • Cost Efficiency Trends:

    • 20062006: $300,000\$300,000

    • 20142014: $1,000\$1,000

    • 20182018: $100\$100 (with tests potentially taking only an hour).

  • Private Consumer Costs (20172017):

    • Paternity Test: $24.99\$24.99 to $125\$125.

    • Ethnicity/Ancestry: $79\$79.

    • DNA Profile: $100\$100 to $2,000\$2,000.