Topic 6.1 Forensic Biology

Topic 6.1 Forensic Biology

1. DNA Profiling

   • A large amount of DNA does not code for proteins.
     • The non-coding blocks are called introns. They are inherited in the same way as any genes within the coding regions (Exons)
       • Within introns, short DNA sequences are repeated many times. = Short tandem repeats (STRs)
         • The same STRs occur at the same place (locus/loci) on both chromosomes of a homologous pair.
           • The number of times they are repeated however can be different.
   • How is a DNA profile made?
     • 1) Obtaining the DNA
       • DNA could come from: Cheek swab, white blood cells in a blood smear, bone marrow in a skeleton or from sperm left after a sexual assault.
       • The tissue sample is physically broken down in a buffer solution that includes salt and a detergent to disrupt the cell membranes
         • The small suspended particles, including the DNA, are separated from the rest of the cell debris by filtering or centrifuging.
           • Protease enzymes remove proteins and then cold ethanol is added to precipitate out the DNA.
     • 2) Creating the fragments
       • Alec Jeffreys - treating the DNA sample with restriction enzymes found naturally in bacteria
         • The enzymes will only cut DNA at specific base sequences (usually 4/6 base pairs long)
           • If the restriction sites are either side of a STR, that fragment of DNA will remain intact but it will be cut away from the rest of the genome
     • 3) Polymerase chain reaction
       • Allow forensics to use tiny deposits of hair, skin or body fluid for identification purposes by copying DNA numerous times.
       • uses DNA primers which are short DNA sequences complementary to the DNA adjacent to the STR.
         • DNA primers are marked with fluorescent tags
         • Placed in a reaction tube with DNA polymerase, DNA primer and nucleotides and undergoes a cycle of temperature changes
           • 1st: separates the double stranded DNA. 2nd: optimises prime binding to the target DNA sequence in the sample. Polymerase attaches and replication occurs. Final temperature: is the optimum temperature for the heat-stable DNA polymerase.
     • 4) Separating the fragments
       • Gel electrophoresis
         • DNA placed on a gel of agarose which provides a stable medium through which the fragments move.
           • The gel is submerged in a buffer solution and connected to electrodes that produce a potential differences across the gel.
             • -ve charged DNA fragments migrate through the gel according to their overall charge and size. Smaller fragments travel faster thus closer to the +ve electrode
               • A reference sample with fragments of known length may be added known as a DNA ladder.
     • 5) Visualising the fragments
       • Southern blotting = Transfer the fragments to a more resilient membrane
         • It is placed directly onto the gel and dry absorbent paper placed on top. This draws buffer solution up through the gel, carrying DNA fragments onto the membrane
           • During this, the fragments are denatured into single strands exposing the base sequence.
             • The membrane is then incubated with an excess of labelled DNA probe (short section of DNA with a base sequence complementary to the target DNA sequence needing to be located.
               • any unbound probe that is not bound to any complementary sequence is washed away.
                 • With a radioactive probe, the membrane is dried and placed next to X-ray film. The film blackens wherever the probe has bound with the DNA to form double-stranded fragments
                 • If the probe is fluorescent, its position on the membrane can be visualised under ultraviolet light.
   • Using the DNA profile
     • Offspring receive one repeated sequence randomly from each parent
       • used for things like identification purposes, settling paternity disputes, identifying stolen animals and looking at variation and evolutionary relationships between organisms

2. Determining time of death

   • As soon as a person dies, a series of physical and chemical changes start to take place in their body.
   • Body Temperature
     • Normal body temperature: 36.2 - 37.6
       • As soon as you die, body temperature drops due to the absence of heat-producing chemical reactions
         • Core body temp is measured via the rectum or through an abdominal stab.
           • Environmental factors must be noted as these will have an impact on how the body has been cooled
             • Follows a sigmoid curve.
               • initial plateau: normally lasts 30-60 mins
     • Useful for estimating time of death during the first 24 hours post-mortem
       • Many factors will affect post-mortem cooling including: body size, body position, clothing, air movement, humidity, temperature of surroundings
   • Rigor mortis
     • Joints become fixed during rigor mortis and their position, whether bent or straight, will depend on the body position at time of death.
       • 1) Muscle cells become starved of oxygen, and oxygen-dependent reactions stop.
         • 2) respiration in the cells becomes anaerobic and produces lactic acid.
           • 3) The pH of the cell falls, inhibiting enzymes and anaerobic respiration
             • 4) ATP needed for muscle contraction is no longer produced. Bonds between muscle proteins become fixed.
               • 5) Proteins can no longer move over one another to shorten the muscle, fixing the muscle and joints.
                 • Rigor mortis passes as muscle tissue starts to break down
                   • Approx time since death?
                     • Warm, not stiff =
                       • Less than 3 hours
                     • Warm, stiff =
                       • 3-8 hours
                     • Cold, stiff =
                       • 8-36 hours
                     • Cold, not stiff =
                       • more than 36-48 hours
       • Smaller muscles stiffen faster than larger ones.
   • Decomposition
     • Tissues starts to break down due to the action of enzymes.
       • Autolysis occurs first: the body's own enzymes begin to break down cells
         • Bacteria from the gut and gaseous exchange system rapidly invade the tissues after death, releasing enzymes
           • Signs of decomposition:
             • Greenish discolouration of the skin of the lower abdomen due to the formation of sulphaemoglobin in the blood. Darken to redish green and then turn to a purple black colour.
               • In average conditions: 36-72 hours after death
             • Gas or liquid blisters may appear on the skin.
             • Gases form in the intestines and tissues causing the body to smell and become bloated.
               • occurs after about a week
         • Environmental temperatures have a major influence: Low temp = slows down High temp = speeds up.
           • The rate of decomposition is highest between 21 and 38'C
             • Intense heat denatures the enzymes involved in autolysis.
   • Forensic Entomology
     • The presence of insects.
       • Samples are taken of any insects found on, near or under the body.
         • Temperature or the air/ground/body and 'maggot mass' are measured
           • Some maggots will be killed at the time of collection so that their age can be determined
             • Some will be taken back to the lab alive and fed on meat, allowing them to complete their development

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