Lecture Notes: Gel Electrophoresis

Introduction

• Gel electrophoresis is a technique for separating DNA or protein molecules based on charge and mass.
• Two types of electrophoresis:
  • Protein electrophoresis: Separates proteins.
  • DNA electrophoresis: Focuses on DNA molecules.
• This lecture focuses on DNA electrophoresis.

DNA Electrophoresis

• Purpose: To separate DNA fragments by charge and mass, producing patterns known as "DNA fingerprints."
• Applications:
  1. Forensics: Identifying individuals at crime scenes using DNA.
  2. Paternity Testing: Confirming biological relationships.
  3. Genetic Screening: Detecting specific alleles.

Requirements for DNA Electrophoresis

1. DNA Fragments:
   • Different lengths and masses, crucial for separation.
   • Example shown: Fragments A (longest) to D (shortest).
2. Polyacrylamide Gel:
   • Used to separate DNA based on mass.
3. Electrical Field:
   • Negative (cathode) and positive (anode) ends are applied.
   • DNA is negatively charged due to phosphate groups.

Process of DNA Electrophoresis

• Setup:

  • Gel is placed in a container with cathode and anode at opposite ends.
  • Wells are created at the cathode end to load DNA samples.
  • Multiple copies of DNA fragments are required for clearer results, achieved through Polymerase Chain Reaction (PCR).

• Separation Mechanism:

  • DNA moves from negative to positive end due to its negative charge.
  • Smaller fragments move faster and farther than larger ones due to less resistance.

• Result Interpretation:

  • Produces bands corresponding to DNA fragments.
  • Each band represents a different DNA fragment size.

Example Application: Genetic Disease Screening

• Gene Variants:

  • Large B allele (normal) not cut by restriction enzymes.
  • Small B allele (disease-causing) can be cut, producing two fragments.

• Case Study:

  • Three individuals tested for genetic variants: Methuselah, Lamia, and Barrett.
  • Results Interpretation:
    • Methuselah: Large B Large B (only one band, not cut).
    • Lamia: Small B Small B (two bands, fragment cut).
    • Barrett: Large B Small B (three bands, mix of cut and uncut).

Conclusion

• Gel electrophoresis is a powerful tool for DNA analysis, useful in various applications including forensic science, paternity testing, and genetic screening.
• Understanding the movement based on charge and mass is critical for interpreting results.