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Molecular biology

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  • DNA/RNA overview
  • Figure 20.9 Gel electrophoresis
  • Transcript

    • 1. Patricia Linton Manchester Metropolitan University
    • 2. It’s just so amazing – that’s what!
    • 3. It’s not science fiction – it’s science fact!
      • DNA - Encodes all of the genetic information needed for the development and functioning of all cells.
      Discipline is full of amazing facts
    • 4. TRANSCRIPTION RNA PROCESSING DNA RNA transcript 3  5  RNA polymerase Poly-A Poly-A RNA transcript (pre-mRNA) Intron Exon NUCLEUS Aminoacyl-tRNA synthetase AMINO ACID ACTIVATION Amino acid tRNA CYTOPLASM Poly-A Growing polypeptide 3  Activated amino acid mRNA TRANSLATION Cap Ribosomal subunits Cap 5  E P A A Anticodon Ribosome Codon E The central dogma DNA RNA PROTEIN
    • 5. TRANSCRIPTION TRANSLATION DNA mRNA Ribosome Polypeptide (a) Bacterial cell Nuclear envelope TRANSCRIPTION RNA PROCESSING Pre-mRNA DNA mRNA TRANSLATION Ribosome Polypeptide (b) Eukaryotic cell
    • 6. Uses of molecular biology – endless – revolutionised science
      • Medical
      • Production of recombinant proteins for treatment of disease
      • Determining the genetic basis of cancer – thereby improving treatment and prognosis
      • Diagnosis of disease – fast diagnosis of HIV/TB – infectious diseases
      • Agricultural
      • pest resistance/ drought resistance/increased productivity
      • Industrial
      • Manufacture of proteins, strain improvement
      • Forensic
      • Used to place criminals at scene of crime or rule out suspects – important for conviction
      • DNA fingerprinting
      • Archaeology
      • Analysis of ancient DNA in ceramic food vessels
      • Environmental Science
      • Identification of species
      • Clean-up of oil spills
    • 7. (a) Tobacco plant expressing a firefly gene (b) Pig expressing a jellyfish gene
    • 8. Kary Mullis – Genius or eccentric? ‘ Science grows like a weed every year’ ‘ Each of us have things and thoughts and descriptions of an amazing universe in our possession that kings in the 17th Century would have gone to war to possess’ ‘ Science consistently produces a new crop of miraculous truths and dazzling devices every year’ ‘ I have had an encounter with an extraterrestrial in the form of a fluorescent raccoon’ ??????? You can read his Nobel lecture here: http://nobelprize.org/nobel_prizes/chemistry/laureates/1993/mullis-lecture.html
    • 9. PCR – in the dark ages 8 BORING hours per PCR! 95º C 5 min 35 times 55º C 3 min 72º C 5 min
    • 10.
      • Individual rows of receptacles can be heated – can try lots of reaction temperatures in one ‘run’
    • 11.  
    • 12. The different steps of PCR
      • Water
      • Buffer
      • DNA template
      • Primers
      • Nucleotides
      • Mg 2+ ions
      • DNA Polymerase
      PCR recipe
    • 13. The temperature profile of a PCR cycle is controlled by the thermal cycler program which results in a near exponential increase in PCR product accumulation for about the first 30 cycles. The 3 Os!!!!!
    • 14.
      • • Used to separate molecules based on their charge and size
      • • Agarose or acrylamide gels can be used to separate DNA fragments
      • • DNA is acidic; it migrates from the negative to the positive
      • end of the gel
      • each fragment’s migration rate is inversely proportional to
      • the log of its molecular
      • weight
    • 15. http://learn.genetics.utah.edu/content/labs/gel/
    • 16. Mixture of DNA mol- ecules of different sizes Power source Longer molecules Shorter molecules Gel Anode Cathode 1 2 Power source – + + –
    • 17.  
    • 18. DNA Fingerprinting
      • Steps in DNA fingerprinting:
        • DNA isolated from tissue sample or PCR’d up
        • DNA cut into fragments with enzymes
          • DNA with different sequences produce fragments of different sizes
          • Can use restriction enzymes to cut either side of VNTRs for genetic typing
        • Fragments separated on basis of size and visualized
        • Each person’s set of fragments is unique
    • 19.  
    • 20.  
    • 21.  
    • 22. DNA Fingerprinting – identification of the remains of the Russian Royal Family
      • DNA fingerprinting showed that 9 different people were buried in the Ekaterinburg grave.
      • Romanovs would be more similar in pattern to each other than to non-relatives.
      • All of a child’s bands must be present in one or both of the parents.
    • 23. 7.6 DNA Fingerprinting Adult 1 Adult 2 Adult 3 Adult 4 Adult 5 Adult 6 Child 1 Child 2 Child 3
    • 24. DNA Fingerprinting
      • To see if parents and their children were Romanovs, DNA fingerprints were prepared for relatives of tsar and tsarina.
      • Adult male skeleton (related to the children) was related to George, the tsar’s brother.
      • Adult female skeleton (related to the children) was related to Prince Philip, the tsarina’s grand-nephew.
      • Conclusion : the grave contained the tsar, tsarina, three of their children, and four servants.
    • 25. Making your own equipment
    • 26.