Gene technologies

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  • Created by: vezting
  • Created on: 07-12-15 10:15
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  • gene technologies
    • PCR (polymerase chain reaction)
      • artificiallDNA replication.
        • generate multiple copies quickly. useful crime scenes
      • 1: reaction mixture- DNA sample, free nucleotides, primers, DNA polymerase.
        • primers complementary to DNA fragment.
      • 2: DNA heated 95dgrs. H bonds break.
      • 3:cooled 65dgrs, primers bind
      • 4: 72dgrs, DNA polymerase lines free nucleotides by template strand. complemantary base pairing.
      • 5:two new copies of fragment DNA made in one round of PCR.
      • rate of PCR is exponential.
      • sequencinf reaction lies upon: anti-parallel sug-phos backbone, strands have 5 prime and 3 prime ends so DNA polymerase knows which way buil, base pairs add according complementary
    • restriction enzymes
      • some sections DNA have palindromic sequences, consist anti-parallel base pairs.
      • restriction enzymes recognise specific palindromic sequences- recognition sequences.
      • DNA cut at sequence. different rest.enzymes cut different seequences, shape complementary active site.
      • if recog.sequence presnt either side useful DNA fragment rest.enzymes isolate.
      • DNA smaple incubated with specific rest.enzymes, cuts DNa by hydrolysis, leaves sticky ends.
        • sticky ends bind fragment to other DNA with complementary sticky ends
          • complementary sticky ends by using same rest.enzyme
    • Gel electrophoresis
      • used to seperate DNa by fragment size
      • 1: DNA + rest.enzymes
        • creates fragments
      • 2:DNA samples placed into wells cut in cathode end of gel.
      • 3:gel immersed into buffer solution, electric current passed through- 2 hours
      • 4:DNA negative due to phosphate groups, DNA diffuse toward annode
      • 5: shorter lengths diffuse quicker and sor move further in the time. position shown by flourescent tags
    • Probes
      • DNA probes recognise specific sequences.
      • isolate genes on chromosomes and locate mutates genes
      • the have specific base sequence complementary tothe DNA, will bind to 'target sequence' if present, identified using flourescent or radioactive tag
      • 1: sample DNA digested to fragments by rest.enzymes, seperated by gel electrophoresis
      • 2:seperated fragments transferred to nylon membrane incubated with flourescent probes
      • 3:membrane exposed to UV light and if target sequence present, will appear flourescent.


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