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Chapter 13. DNA: The Indispensable Forensic Science Tool. Objectives:. Describe and demonstrate DNA collection, preservation, and testing, using gel electrophoresis. Use Product Rule to quantify and interpret results. Father of DNA fingerprinting. Alec Jeffreys 1985

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chapter 13
Chapter 13

DNA: The Indispensable Forensic Science Tool

objectives
Objectives:
  • Describe and demonstrate DNA collection, preservation, and testing, using gel electrophoresis.
  • Use Product Rule to quantify and interpret results.
father of dna fingerprinting
Father of DNA fingerprinting
  • Alec Jeffreys
  • 1985
  • Responsible for the revelation of DNA fingerprinting
dna fingerprinting
DNA fingerprinting
  • Isolating and reading DNA markers (portions of the DNA structure of certain genes that are unique to each individual).
what is a gene
What is a gene?
  • Fundamental unit of heredity.
  • Instructs body cells to make proteins.
  • The proteins are responsible for determining specific traits.
what is dna
What is DNA?
  • Polymer that carries the body’s genetic information.
  • Double helix.
  • Watson & Crick – built ball & stick model of DNA.
what is a polymer
What is a polymer?
  • Large molecule that links together a series of repeating units.
  • In DNA, the repeating units are called nucleotides.
what are nucleotides
What are nucleotides?
  • Building blocks of DNA molecule.
  • Composed of
    • Sugar (backbone)
    • Phosphate group (backbone)
    • Nitrogen base.
4 nitrogen bases in dna
4 Nitrogen Bases in DNA
  • Adenine + Thymine = complementary bases.
  • Cytosine + Guanine = complementary bases
  • Three nitrogen bases are called a codon.
  • Each codon codes for a specific amino acid.
  • Groups of amino acids produce a protein.
dna to protein
DNA to Protein
  • The specific proteins produced by a cell are directly related to the sequence of nucleotides in the DNA of the cell.
  • The individuality of an organism is determined by the organism’s DNA nucleotide sequence.
  • Sickle cell anemia (example)- abnormal hemoglobin
human genome
Human Genome
  • Total DNA content found within the nucleus of a cell.
human genome project
Human Genome Project
  • Global effort designed to reveal the location of all genes in the human genome.
  • Will be useful in diagnosing & treating genetic diseases.
  • Will help reveal the role & implications of evolution.
dna replication
DNA Replication
  • Synthesis of new DNA from existing DNA.
  • Semiconservative.
steps of dna replication
Steps of DNA Replication
  • Helicase
    • Enzyme that holds double strands apart (forks)
  • DNA Primer
    • Attracts DNA polymerase
  • DNA Polymerase
    • Enzyme that inserts complementary base pair & proofreads the new DNA strand for errors.
  • Ligase
    • Enzyme that seals the new strand to the old strand of DNA.
slide15
PCR
  • Polymerase Chain Reaction.
  • Technique for copying a particular DNA portion of interest outside a living cell.
  • Can produce many exact copies of segments of DNA so sample size is no longer a limitation of DNA physical evidence.
  • 1st step is to heat the DNA strands to cause them to separate completely.
  • Each cycle of the DNA Thermal Cycler takes approximately 2 minutes.
recombinant dna
Recombinant DNA
  • The cutting of DNA into fragments that can later be incorporated into another DNA strand.
  • Enormous commercial implications (ex: growth hormones, insulin, etc)
restriction enzymes
Restriction Enzymes
  • Chemicals that act as scissors to cut DNA molecules at specific locations.
  • Produces RFLPs.
rflps
RFLPs
  • Restriction Fragment Length Polymorphism
  • Different fragment lengths of base pairs that result from cutting a DNA molecule.
  • All humans have the same type of repeats, but vary in the number.
  • Typically a core repeat = 15-35 bases.
  • Radioactive probes are used to visualize RFLPs (make them gel visible).
tandem repeats
Tandem Repeats
  • Region of a chromosome that contains multiple copies of a core DNA sequence that are arranged in a repeating fashion.
  • Origin & significance is a mystery.
  • About 30% of human genome is largely composed of repeating segments of DNA.
  • It is thought that they may act as spacers between the coded regions of DNA.
steps of dna typing
Steps of DNA Typing
  • Digestion with restriction enzymes.
  • Electrophoresis
  • Southern blotting
  • Hybridization with a radioactive probe
  • Process with X-ray film
  • In a three probe system of DNA typing, the typical DNA pattern will show a maximum of about 6 bands.
  • One band can indicate that the individuals are probably homozygous for the specific gene.
gel electrophoresis
Gel Electrophoresis
  • DNA is separated by fragment size.
  • Smaller size fragments move faster than larger fragments.
southern blotting
Southern Blotting
  • The transfer of DNA fragments onto a nylon membrane.
hybridization
Hybridization
  • Process of joining two complementary strands of DNA to form a double stranded molecule.
gene splicing
Gene Splicing
  • Laboratory procedure responsible for making possible the development of bacteria that can synthesize insulin.
advantages of working with short dna fragments
Advantages of Working With Short DNA Fragments
  • They are more stable & less likely to break apart.
  • They are less subject to degradation due to adverse environmental conditions.
  • Quantity can be greatly amplified by PCR technology.
slide26
STR
  • Short Tandem Repeats
  • Region of a DNA molecule that contains short segments consisting of 3 -7 repeating base pairs.
  • Most successful & widely used DNA profiling process.
  • Commercial STR kits include means to detect the amelogenin gene to allow for gender determination.
str vs rflp
STR vs RFLP
  • STR analysis has replaced RFLP DNA typing because:
    • It is less subject to sample degradation.
    • It reduces the time to obtain results from a sample.
    • It requires a smaller sample size.
str capillary electrophoresis
STR Capillary Electrophoresis
  • Evolved from flat gel electrophoresis.
  • Automates sampling and data collection.
  • Decreases analysis time.
multiplexing
Multiplexing
  • The concept of simultaneously extracting, amplifying, and detecting a combination of STRs.
  • Y-STR markers are useful in cases where multiple males are involved in a sexual assault on a victim. (expect 3 peaks).
mtdna
mtDNA
  • Mitochondrial DNA.
  • HV1 & HV2 are regions of mtDNA.
  • Few forensic labs do mtDNA analysis because
    • Analysis procedure is very rigorous.
    • Costs much more than nuclear DNA profiling.
    • Study takes time.
    • Discriminating power of mtDNA is less than that of STR analysis.
dna profiling technology
DNA-profiling technology
  • Has been used to
    • Settle matters of questioned paternity/maternity.
    • Match suspect(s) to biological evidence found at a crime scene.
    • Decide immigration cases based on family relationships.
codis
CODIS
  • Shared databases of DNA typing information from convicted felons and crime scene evidence.
luminol
Luminol
  • Detects traces of blood without compromising potential DNA typing.
buccal cells
Buccal Cells
  • Cells obtained from the mouth and inside of the cheek.