DNA: The Indispensable Forensic Science Tool

1. DNA: The Indispensable Forensic Science Tool Chapter 9

2. Introduction • 1985-Sir Alex Jeffreys recognized that portions of DNA structure of certain genes are unique to each individual. • The process of identifying these markers is known as DNA fingerpringting, DNA typing, or DNA profiling • Forensic Scientists now had a way of linking biological samples like blood, semen, hair, and tissue to a single individual • US courts have accepted the reliability of DNA evidence

3. DNA = Deoxyribonucleic Acid • DNA is a polymer • Polymers are very large molecules put together by linking units called monomers • The monomers of DNA are called nucleotides.

4. Nucleotides • DNA nucleotides consist of : • A sugar molecule that forms the backbone • A phosphate group which links the sugars • A nitrogen containing base (adenine, cystine, guanine, thymine) • Nucleotides are linked together to form a double helix What is DNA?

5. Complementary Base Pairing • Adenine always pairs with Thymine • Guanine always pairs with Cytosine • Bases are joined together by hydrogen bonding.

6. How DNA works • DNA directs the production of proteins which play a basic role in the structure and function of living organisms • Proteins are built by linking amino acids • The are 20 amino acids that make up proteins • The sequence of amino acids determines the shape and function of the protein

7. How DNA works • Each amino acid is coded by a sequence of 3 bases

8. Replication of DNA • Replication – synthesis of new DNA from existing DNA • The double helix is unwound and a copy is made. • DNA polymerase – • Assemble new DNA strands in the proper base sequence • Correct mismatched base pairs

9. Polymerase Chain Reaction (PCR) • A technique for replicating a portion of DNA outside of a living cell • DNA polymerases are used to make copies of DNA material • This is useful to forensic scientists because small samples could be multiplied.

10. DNA Typing with Tandem Repeats • Tandem repeat –sequence of base pairs repeated numerous times. • ~30 % of human genome is made up of tandem repeats. • They act as spacers in between coding regions of the DNA • Used by forensic scientists to distinguish individuals.

11. DNA Typing with Tandem Repeats • Humans have the same type of repeats but a wide variety in the number of repeats. • Restriction fragment length polymorphisms (RFLP’s) – repeat segments cut out of DNA by restriction enzymes.

12. DNA Typing with Tandem Repeats • Length differences associated with RFLP’s allow forensic scientists to identify individuals. • Electrophoresis is used to separate different sized fragments. • Gel Electrophoresis

13. DNA Typing with Tandem Repeats

14. Polymerase Chain Reaction (PCR) • PCR replaced RFLP techniques in the mid-1990’s The Basics: • Short DNA sequences are identified. • DNA strands are unwound and replicated yielding two copies of the original sample • 28-32 cycles are carried out yielding over a billion copies of the DNA strand. Polymerase Chain Reaction

15. Polymerase Chain Reaction (PCR) Advantages of PCR over RFLP typing: • PCR uses much smaller strands of DNA • Smaller strands are more stable and less subject to degradation • Smaller strands overcome the sample-size problem associated with crime-scene evidence

16. Short Tandem Repeats (STR’s) • STR’s – DNA profiling procedure used currently • Short Tandem Repeats – regions of DNA molecules containing short segments of 3 to 7 repeating base pairs. • STR’s are less susceptible to degradation and can be amplified by PCR • Multiplexing: A technique that simultaneously detects more than one DNA marker in a single analysis.

17. Short Tandem Repeats (STR’s) • In the US, the forensic science community has standardized 13 STR’s for entry into the CODIS database • FBI Core STR Loci • Calculating the frequency of occurence of a DNA profile • Separation of STR's by capillary electrophoresis • Results of electrophoresis on STR's

18. Sex Identification Using STR’s • The sex of a DNA contributor can be determined by looking at the amelogenin gene. The gene is 6 bases shorter on the X chromosome than it is on the Y chromosome. • Y-STR’s – STR’s found on the Y chromosome; useful when there is a mix of DNA from more than one male.

19. Mitochondrial DNA • Mitochondrial DNA (mtDNA) – found in the mitochondria (outside of the cell nucleus) • mtDNA is inherited solely from the mother • Forensic Scientists use mtDNA to identify subjects when nuclear DNA is not available. • Frequencies of sequences in the FBI database are about 1 % • inheritance of mtDNA • structure of mtDNA

20. The Combined DNA Index System (CODIS) • CODIS – software program and database developed by the FBI • Contains DNA profiles of convicted offenders, unsolved crime-scene evidence, and profiles of missing people.

21. Collection and Preservation of DNA Evidence • STR profiling requires only 125 picograms of DNA • One human cell contains ~ 7 pg of DNA • Only 18 cells are necessary • As a result, DNA has been successfully analyzed from stamps, envelopes, cups, cigarette butts, chewing gum, skin cells, etc.

22. Collection and Preservation of DNA Evidence • Biological evidence should be documented by notes, sketches, and photographs first • The shape and position of bloodstains should be investigated prior to collection. • Latex gloves will protect investigators from infection AND avoid contaminating the evidence. • Items like clothing should be collected from suspects and victims even if blood isn’t immediately visible.

23. Collection and Preservation of DNA Evidence Packaging of Evidence: • Biological evidence should not be packaged in an airtight container; paper bags, envelopes, or cardboard boxes can be used. Samples should be refrigerated and kept out of direct sunlight • Blood stains can be removed from a surface using a cotton swab soaked in distilled water. • Swabs must be dried prior to packaging

24. Collection and Preservation of DNA Evidence Obtaining DNA Reference Samples: • Reference samples can be obtained by collecting blood or a buccal swab from an individual • If the individual is not available: • Tootbrush • Combs and hairbrushes • Razor • Dirty laundry • Cigarette butts

25. Collection and Preservation of DNA Evidence To minimize contamination: • Change gloves before handling each new piece of evidence • Collect a substrate control. • Use forceps to pick up small items • Package items is individual well-ventilated containers. • Luminol does not destroy DNA