Chapter 12 DNA and RNA - PowerPoint

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Chapter 12 DNA and RNA - PowerPoint Powered By Docstoc
					              Chapter 9
    DNA The Indispensible Forensic
            Science Tool
     Learn the parts of a nucleotide and know how they are linked together to make
                                               DNA.
                  Understand how DNA strands coil into a double helix.
     Describe the concept of base pairing as it relates to the double helix structure of
                                               DNA.
    Explain how the sequence of bases along a DNA strand ultimately determines the
                   structure of proteins that are synthesized within the body.
    Describe how a double-strand DNA replicates itself. What are the implications of
                                this process for forensic science?


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DNA




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              The Structure of DNA
            • DNA is composed of nucleotides
            • Each DNA nucleotide has:
              – Sugar – deoxyribose
              – Phosphate Group
              – Nitrogen Base
                • Purines – double ring – adenine & guanine
                • Pyrimidines – single ring – thymine &
                  cytosine


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                  DNA Nucleotides
      • Chargaff’s
        Rule
            – % of guanine
              always
              equals
              amount of
              cytosine
            – % of adenine
              always
              equals
              amount of
              thymine
            – G=C
            – A=T
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                      X-Ray Evidence
     • Rosalind Franklin
            – used x-ray crystallography
              to photograph DNA
            – Photograph 51 used by
              Watson and Crick to build
              model
            – Visit these links to find out
              more about Miss Franklin
              and her photo
              http://www.pbs.org/wgbh/nova/photo51/
              http://www.providencepictures.com/pp/pre
              ss_photo51_globeandmail.html



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            Watson and Crick Model
      • Shape is a “twisted
        ladder” or Double Helix
            – Sides - sugar and
              phosphates
            – Steps – nitrogen base
              pairs
                • adenine and thymine
                  OR guanine and
                  cytosine
            – Link for interactive
              tutorial:
                • http://www.mrc-
                  lmb.cam.ac.uk/DNA/ind
                  ex.html
                • To use site download
                  Chime Plug-In from mr-
                  ippolito-science-
                  class.org


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            Chromosomes and DNA
                 Replication




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            DNA and Chromosomes
        • Chromosomes are made of DNA &
          Proteins
            – DNA is wrapped around the protein histone
            – Nucleosomes are groups of histones




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                       DNA Replication
            •       DNA makes a copy of itself before cell
                    division
                  1. DNA separates into two strands
                  2. Each strand acts as a template
                  3. Complementary strands formed by base
                     pairing
                  4. Catalyzed by the enzyme DNA polymerase
            visit these links:
            http://academy.d20.co.edu/kadets/lundberg/DNA_animations/DNAreplication.mov
                    http://academy.d20.co.edu/kadets/lundberg/DNA_animations/dna.dcr




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                               DNA Unzips
                                    This will
                                    a special
                                    The enzyme
                                    It does this
                                    until the
                                    slowly
                                    by breaking
                                    enzyme opens
                                    DNA exposed
                                    allow molecule
                                    the double
                                    bases to down
                                        weak
                                    proceedsbe
                                    is completed
                                    hydrogen of
                                    helix
                                    matched
                                    the middle
                                    opened
                                    the DNAnew
                                    bonds between
                                    forming
                                    “sides”
                                    molecule
                                    bases
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            DNA Polymerase
                                                    The enzyme
                                                    Each strand
                                                    New
                                                    Nucleotides
                                                    Resulting
                                                    positions them
                                                    is half new
                                                    hydrogen
                                                    from cell
                                                    in two the
                                                    opposite old
                                                    and half
                                                    storageform
                                                    bonds area
                                                    identical to
                                                    proper base in
                                                    are brought
                                                    DNA of
                                                    between the
                                                    strands base
                                                    form new
                                                    by enzymes
                                                    bases
                                                    DNA
                                                    pairs

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       RNA and Protein Synthesis




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                 Structure of RNA
       • Each RNA nucleotide has:
            – Sugar –ribose
            – Phosphate Group
            – Nitrogen Base
               • Purines – double ring – adenine & guanine
               • Pyrimidines – single ring – uracil & cytosine




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                 Types of RNA
• Three kinds of RNA
     – Messenger RNA (mRNA) – carries instructions to
       ribosome
     – Transfer RNA (tRNA) – carries amino acids to
       ribosome
     – Ribosomal RNA (rRNA) – forms part of the ribosome




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                      Transcription
            •   DNA copies its nucleotide sequence into
                RNA
                1. RNA polymerase binds to DNA promoter
                   region.
                2. DNA separates into two strands.
                3. RNA polymerase uses one DNA strand as a
                   template.
                4. Complimentary bases match template to
                   form RNA.
                5. RNA leaves and DNA closes




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                   RNA Editing
        • Introns – parts of
          DNA that form
          pre-edited RNA
          that are cut out of
          the mRNA
        • Exons – parts of
          DNA that form
          parts of pre-
          edited RNA that
          are spliced
          together to make
          mRNA.

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                    Genetic Code
       • Nitrogen base
         sequence “names”
         amino acids of a
         protein
            – 20 different amino
              acids
            – 4 bases A C G U
            – 3 bases names an
              amino acid (codon)




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                                     Translation
       •        mRNA decoded at ribosome to form
                proteins
            1.       mRNA attaches to ribosome
            2.       tRNA anticodon matches mRNA codons
            3.       Amino acids on tRNA’s bond together
            4.       Ribosome moves to next repeating process
            5.       At end mRNA, protein and ribosome released
       •        Visit: Lew-Port’s Biology Place - Protein Synthesis Animation
            –        Biostudio Protein Synthesis Animation




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      Polymerase Chain Reaction
        • lab technique used to “amplify” small
          amounts
            – uses Taq polymerase derived from hot
              springs bacteria
            – in a few hours 30 cycles results billion
              fold increase
            – for more information visit:
               • http://www.sumanasinc.com/webcontent/anis
                 amples/molecularbiology/pcr.html

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        Recombinant DNA (rDNA)
      • inserts genomic DNA from one species
        into bacterial plasmids
            – uses restriction enzymes to remove
              specialized sections of DNA
            – once in bacterium “product” coded for by
              DNA can be identified
              • used to produce
                 – human growth hormone
                 – insulin


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            DNA Tandem Repeats
        • DNA sequences that are found in
          multiple copies in human DNA
            – seem to act as “fillers” or “spaces” between
              coding regions
            – allow forensic scientists to distinguish
              individuals through DNA Typing




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            Restriction Fragment Length
                   Polymorphisms
        • are repeating DNA sequences that can be
          removed by restriction enzymes
             – length differences in these sequences can be
               used to distinguish individuals
             – sequences are separated by gel electrophoresis
                • Southern blotting transfers separated fragments to
                  nylon membrane
                • Hybridization attaches radioactive compliments to
                  strands
                • Radioactive material “develops” film to form “picture”
                  of DNA fragments


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            Short Tandem Repeats
       • newer DNA profiling technique
            – needs less DNA because sequences are smaller
            – multiple repeats of same bases
               • THO1 – A-A-T-G repeats
            – PCR amplifies the STRs
       • Multiplexing
            – allows multiple different STRs to be identified
              allowing “narrowing” identification in population
            – 13 different CODIS STRs are found with different
              frequencies within the population

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            Capillary Electrophoresis
      • allows automation /
        shortens time
            – thin glass column
              coated with gel
              immersed in buffer
            – DNA injected in one
              end
            – as STRs move
              through gel a
              detector tracks
              them to form a
              electropherogram




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      Sex Identification with STRs
            • amelogenin gene
              – controls formation of tooth pulp
              – on both the X chromosome and Y
                chromosome
                • X version six (6) bases shorter than Y
                  version
                   – females with XX genotype show one band
                   – males with XY genotype show two bands
                • 20 different Y-STR markers identified
                   – can distinguish between males in sexual
                     assaults

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              Mitochondrial DNA
            • Mitochondria
              – powerhouse of cell – site of aerobic respiration
              – has own DNA – mtDNA
              – only maternal DNA
            • mtDNA – circular chromosome of some
              17,000 bases – HV1 and HV2 regions
              used
              – used when no nuclear DNA available
              – used to ID remains (skeletal)
              – need reference samples from female family
                members
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Collect Biological Evidence for DNA
              Analysis
            • STR profiling allows typing with as few
              as 125 picograms of DNA
              – picogram = 10-12 gram or .000000000001
                gram
            • 7 picograms of DNA in a human cell
              – as little as 18 cells can allow a valid STR
                profile to be developed
              – goal to get profile from ONE (1) human
                cell
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            Common Sources of DNA




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                 Proper Collection
            • DNA is in body fluids – infectious
              – disposable gloves
                 • change often
              – face masks
              – shoe covers
              – overalls
            • must prevent “cross contamination”


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  Packaging Biological Evidence
        • moisture grows bacteria and
          mold
            – always in paper or well
              ventilated box
            – swabs need to be air-dried for
              5-10 minutes before boxing
            – cool storage out of sunlight
               • exception: blood in soil must be
                 FROZEN in glass/plastic
        • substrate control
            – sample of unstained surface
              material for comparison
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      Reference DNA Specimens
            • DNA in samples compared to victims
              and suspects
              – 7 ml blood collected with EDTA (inhibits
                enzyme degradation) in sterile vacuum
                tubes and refrigerated
              – buccal swabs – cheek cells for sample




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Contamination of DNA Evidence
        •   as simple as a cough or a sneeze
            – prevention
              1. change gloves with each sample
              2. substrate control for each sample
              3. disposable forceps for each small item
                 (cigarette butts, licked stamps, drinking
                 straws, etc.
              4. each in own ventilated container



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