LESSON 9: DNA Fingerprinting

• DNA profiling - technique used by scientists to distinguish between individuals of the same species using only samples of their DNA
• Alec Jeffreys - invented the process of DNA fingerprinting at the University of Leicester in 1985

  • he was knighted (Most Excellent Order of the British Empire) in 1994
• DNA Polymorphism

  Tandem Repeats

  • satellites
  • minisatellites
  • microsatellites

  Single Nucleotide Polymorphism

  • single-base substitutions
  • single-base insertion/deletions
• sequence polymorphism - single nucleotide polymorphism

  
• length polymorphism - tandem repeat

  
• Minisatellites - Variable Number of Tandem Repeats (VNTR)

  • 9-24 bp repeats
• Microsatellites - Short Tandem Repeats (STR)

  • 2-6 bp repeats
• minisatellites (VNTR)

  
• microsatellites (STR)

  
• DNA fingerprinting - to identify various components of DNA, unique for an individual

  • technique is based on mini-satellites, repeated several times in the genome
• probe - synthesis from minisatellite and labelled with isotopes which can bind onto each minisatellite for visualization
• minisatellites - segments that vary in size and composition and have no apparent function
• DNA Fingerprinting using VNTRs

  • on some human chromosomes, a short sequence of DNA has been repeated a number of times, which may vary from 1-30 repeats
  • these repeat regions are usually bounded by specific restriction enzyme sites
  • cut out the segment of the chromosome containing this variable number of tandem repeats (VNTRs)
  • identify the VNTRs for the DNA sequence of the repeat
• Stages of DNA Profiling

  Stage 1: Cells are broken down to release DNA

  • polymerase chain reaction (PCR) - amplifying available DNA if it is only a small amount
• Stages of DNA Profiling

  Step 2: The DNA is cut into fragments using restriction enzymes

  • each restriction enzyme cuts DNA at a specific base sequence
• restriction fragments - sections of DNA that are cut out
• restriction enzymes - also called restriction endonuclease

  • 1962: "molecular scissors" discovered in bacteria
• restriction enzymes - an enzymatic immune system that recognizes and destroys foreign DNA
• palindromic sequence - one in which the 5' to 3' base is identical in both strands
• Enzyme Site Recognition

  1. each enzyme digests (cuts) DNA at a specific sequence = restriction site
  2. enzymes recognize 4- or 6- base pair, palindromic sequences

  
• restriction endonucleases - named for bacterial genus, species, strain, and type
• enzyme: SmaI

  source: Serratia marcescens, 1st enzyme

  recognition sequence: CCCGGG
• enzyme: HaeIII

  source: Hemophilus aegyptius, 3rd enzyme

  recognition sequence: GGCC
• enzyme: HindII

  source: Hemophilus influenzae, strain d, 2nd enzyme

  recognition sequence: GTPyPuAC
• enzyme: HindIII

  source: Hemophilus influenzae, strain d, 3rd enzyme

  recognition sequence: AAGCTT
• enzyme: BamHI

  source: Bacillus amyloliquefaciens, strain H, 1st enzyme

  recognition sequence: GGATCC
• Stages of DNA Profiling

  Stage 3: Fragments are separated on the basis of size using a process called gel electrophoresis

  • DNA fragments are injected into wells and a current is applied along the gel
  • DNA is negatively charged so it is attracted to the positive end of the gel
  • the shorter DNA fragments move faster than the longer fragments
  • DNA is separated on basis of size
  • radioactive material is added to produce fluorescent image
  • photographic copy of the DNA bands is obtained
• Stages of DNA Profiling

  Stage 4: The pattern of fragment distribution is then analyzed
• biological material used for DNA profiling

  1. blood
  2. hair
  3. saliva
  4. semen
  5. body tissue cells
  6. DNA samples from vaginal cells transferred to the outside of a condom
• Practical Applications of DNA Fingerprinting

  1. Paternity and Maternity
  2. Criminal Identification and Forensics
  3. Diagnosis of Inherited Disorders
• forensic science - use of scientific knowledge in legal situations

  • the chances of two people having exactly the same DNA profile is 30,000 million to 1 (except for identical twins)
• paternity and maternity information can be used in

  1. paternity suits
  2. inheritance cases
  3. immigration cases