DNA Profiling

DNA Profiling

Introduction

  • DNA profiling, also known as DNA fingerprinting, is a process of matching an unknown DNA sample to a known DNA sample.
  • This is useful in crime scenes to identify victims and murderers.
Scenario: Crime Scene
  • A crime scene is depicted with a victim (Georgie) and potential evidence like blood.
  • The goal is to identify the murderer.
  • Traditional methods like fingerprinting may not always be viable (e.g., if the perpetrator wiped everything clean).
  • If there are blood samples at a crime scene that contains cells, scientists can extract DNA from those blood.
Identifying Suspects
  • A witness identifies three potential suspects: Ronaldo, Johnny Bravo, and John Wick.
  • The challenge is to determine which suspect's DNA matches the unknown sample from the crime scene.

DNA Profiling Explained

  • The process involves comparing the unknown DNA sample from the crime scene to known samples from the suspects.
  • If there's a blood splatter, cells with DNA are extracted.
  • The DNA is not enough from the blood splatter so DNA needs to be replicated and amplified.

PCR: Polymerase Chain Reaction

  • PCR is a laboratory technique used to amplify a small amount of DNA into a large amount of DNA, creating many copies.
Process of PCR
  1. DNA Extraction:
    • A segment of DNA is extracted.
  2. Test Tube Preparation:
    • The extracted DNA is placed in a test tube with primers, free nucleotides (A, C, T, G), and Taq DNA polymerase.
      • Primers serve as a starting point for DNA replication.
      • Free nucleotides are the building blocks for the new DNA strands.
      • Taq DNA polymerase is a heat-resistant enzyme that adds nucleotides to create the new DNA strands.
  3. Thermocycler:
    • The test tube is placed in a thermocycler, which changes temperatures to facilitate DNA replication.
Steps in PCR
  1. Denaturation:
    • The thermocycler heats to 95°C to break the hydrogen bonds between DNA strands, separating them.
  2. Annealing:
    • The temperature is lowered to 55°C, allowing primers to attach to the separated DNA strands, acting as a starting point for replication.
  3. Extension:
    • The temperature is raised to 75°C, and Taq DNA polymerase adds free nucleotides to the primers, extending the DNA strands and creating new, complementary strands.
PCR Results
  • After one cycle, one copy of DNA becomes two.
  • Each cycle doubles the amount of DNA:
    • 1 cycle: 21=22^1 = 2 copies
    • 2 cycles: 22=42^2 = 4 copies
    • 3 cycles: 23=82^3 = 8 copies
    • n cycles: 2n2^n copies
  • PCR amplifies DNA, creating billions of copies in hours.
Why is it Called PCR?
  • Polymerase: DNA polymerase is used to synthesize new DNA.
  • Chain Reaction: Process involves multiple cycles of replication.

Gel Electrophoresis

  • Gel electrophoresis is a laboratory technique used to separate molecules (DNA, RNA, proteins) based on size and charge.
    Specifically, it's a lab technique that separates molecules according to size and other properties like charge.
Process of Gel Electrophoresis
  1. Enzyme Cutting:
    • DNA samples from suspects are treated with an enzyme that cuts DNA at specific sequences.
    • The cuts result in DNA fragments of different sizes.
  2. Gel Preparation:
    • A gel is prepared with small wells for DNA samples.
    • The gel is placed in an electrophoresis machine with positive and negative electrodes.
  3. Sample Loading:
    • Cut DNA fragments from each suspect and the crime scene are placed in separate wells.
  4. Electrophoresis:
    • An electric current is applied, causing the negatively charged DNA fragments to move towards the positive electrode.
    • Smaller DNA fragments move faster and further through the gel, while larger fragments move slower and less far.
  5. Visualization:
    • The separated DNA fragments create a pattern of bands in the gel.
    • The pattern is visualized using staining or other techniques.
Gel Properties
  • The gel contains a mesh-like structure with tiny fibers.
Movement of DNA Fragments

  • Big pieces: Struggle to move and thus travel less distance.

  • Small pieces: Travel further because they can pass through the mesh more easily.

  • Size/weight: Heavy pieces travel less far; lighter pieces travel further.

Results and Interpretation
  • The DNA from the crime scene sample is compared to the DNA from the suspects.
  • The suspect whose DNA pattern matches the crime scene DNA is identified as the likely perpetrator.
  • In the example, John Wick's DNA matches the crime scene DNA.

DNA Profiling Applications

  • Crime Investigation: to prove who is guilty.
  • Parentage: Determine parentage through DNA matching since people in the same family will have similar results.
  • Evolutionary Relationships: Comparing DNA similarity between species to understand evolutionary connections.

Summary Steps

  1. Collect samples from Crime Scene and Suspects.
  2. DNA Extraction: Extract DNA from the collected samples.
  3. Amplify: PCR is used to make more copies.
  4. Gel Electrophoresis: Cuts the extracted samples with the same enzyme and runs PCR samples through the gel to create bands.
  5. Analyze and Interpret: Examine the gel electrophoresis and analyze the band patterns to see who the murderer is.

Quick Questions

  1. A small amount of suspects DNA is obtained from a crime scene. What techniques would be used to carry out DNA profiling?
    • Answer: PCR and Gel Electrophoresis
  2. What is the reason for Taq DNA polymerase being used in the polymerase chain reaction or PCR?
    • Answer: Because it can function at very high temperatures.
  3. The diagram shows results of electrophoresis of DNA from a crime scene. Which suspects would be implicated as the criminal according to the gel of DNA shown?
    • Answer: Suspect Four