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DNA Fingerprinting. Use of DNA to Determine Identity. DNA controls production of proteins Results in phenotype (eye color, facial features) Contributes to structure and function DNA – 3 billion base pairs No other person on planet has same code (except identical twins!)

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use of dna to determine identity
Use of DNA to Determine Identity
  • DNA controls production of proteins
    • Results in phenotype (eye color, facial features)
    • Contributes to structure and function
  • DNA – 3 billion base pairs
    • No other person on planet has same code (except identical twins!)
  • Only 0.1% of DNA differs from person to person
    • But the regions that do vary provide a true genetic blueprint
use of dna to determine identity1
Use of DNA to Determine Identity
  • DNA obtained from skeletal remains
  • DNA purified
  • If degraded – amplified w/ PCR
  • RFLPs compared to determine final identity of missing person
what are some sources of dna evidence
What are some sources of DNA Evidence?
  • Skin cells
  • Hair
  • Blood
  • Semen
  • Anything with DNA
dna fingerprinting1
DNA Fingerprinting
  • AKA - DNA profiling analysis or DNA typing
  • Electrophoretic analysis of DNA fragment sizes generated by restriction enzymes
  • Provides accurate, unambiguous identification of source DNA samples
restriction enzymes
Restriction Enzymes
  • Endonucleases that cut phosphate bonds
    • Break bonds between deoxyribose and phosphate
  • Attach to DNA and “read” nucleotides
  • Cut at specific sequences
  • Over 3000 types
restriction enzymes1
Restriction Enzymes
  • Named after the organisms from which they were discovered / isolated
  • Eco RI – Escherichia coli RY13
  • Hind III – Haemophilus influenzae R4
  • Bam HI – Bacillus amyloliquefaciens H
eco ri function
Eco RI Function

Eco RI

5` - G A A T T C – 3`

3` - C T T A A G – 5`

5`- G A A T T C – 3`

3` - C T T A A G – 5`

DNA fragments with “sticky” ends

restriction enzymes2
Restriction Enzymes
  • Forensic labs – use at least 2 restriction enzymes
    • 4-base and 5-base (sometimes others)
  • Size of DNA fragments – depends on distance between recognition sites
  • Longer DNA molecule – the greater probability that a specific recognition site will occur
restriction enzymes3
Restriction Enzymes
  • Average human chromosome = 100 million bp
  • Eco RI – 6 bp recognition site
  • Probability – 1 site / 4096 bp
  • Cut human DNA into ~25,000 fragments
restriction enzymes4
Restriction Enzymes
  • No two individuals have same pattern of restriction enzyme recognition sites
  • Each person – unique genotype (different alleles)
  • Mutations / Insertions / Deletions
    • Changes distribution and frequency of restriction enzyme recognition sites
after restriction digest
After Restriction Digest
  • Analyze DNA fragments on agarose gel
  • DNA fragments separated by molecular weight (size) due to net negative charge on phosphates
  • Restriction enzyme cleavage of relatively small DNA molecules – key to RFLP analysis
  • If DNA fragments are too large – can’t see RFLP pattern
sample rflp patterns
Sample RFLP Patterns

Suspect RFLP #4

Control RFLP

Suspect RFLP #1

Suspect RFLP #3

Suspect RFLP #5

Suspect RFLP #6

Molecular Marker

Suspect RFLP #2

restriction enzyme digestion
Restriction Enzyme Digestion
  • Be sure to label all reaction tubes (4)
    • 10 μl of enzyme reaction buffer
    • 15 μl of DNA sample from missing person
    • 15 μl of Restriction Enzyme
  • Incubate at 37°C for ~60 minutes
  • Add 5 μl of Gel Loading Solution after incubation
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