Genetic Engineering and Recombinant DNA Technology

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Genetic Engineering and Recombinant DNA Technology

• Diversity of life is based on the differences in
  genes (DNA)
• 1 gene 1 protein 1 function?
• If we change the DNA (genes), then it would be
  possible to change the function of the cell, and
  ultimately the organism

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Genetic Engineering and Recombinant DNA Technology

• Recombinant DNA-isolating and combining DNA of
  one organism with the DNA of a different organism
• Human Insulin gene ? bacteria
• Genetic Engineering-modifying gene(s) to benefit
  or cure an organism
• Agriculture? crops, animals
• Gene therapy when cloned genes are used to
  modify humans, to control genetic diseases ?
  sickle cell, hypercholesterolemia

Ice cream anyone?
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GloFish
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Cloning Genes

• Cloning-production of genetically identical DNA
• To clone a gene, must combine human DNA with
  another organisms DNA ? recombinant DNA
• Put gene into a plasmid ? extra bacterial
  chromosome that can accept foreign DNA

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Molecular Tools Used For Recombinant DNA

• How to combine the DNA from 2 different
  organisms?
• DNA must be cut and put back together
• What kind of molecule could cut DNA and put it
  back together again?

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Molecular Tools Used For Recombinant DNA
Restriction Enzymes

• Bacterial enzymes that cut DNA at specific DNA
  sequences
• Leaves single stranded tails at the end of the
  DNA fragments ? sticky ends
• DNA fragments cut by same enzyme will come
  together and form a 2x strand DNA fragment
  sticky ends are complementary
• DNA ligase closes the nicks in the DNA

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Molecular Tools Used For Recombinant DNA
Polymerase Chain Reaction (PCR)

• Multiple copying of a specific DNA sequence
• Requires
• primers
• Heat insensitive DNA polymerase from Thermus
  aquaticus (Taq)
• Used when only have small amounts of DNA and need
  to do many different tests genetic identification

Cycle 30 5.37 x 108
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Genetic Fingerprinting

• Every person has specific DNA sequences that are
  unique like fingerprints
• During a criminal investigation, those are the
  sequences that are studied using restriction
  enzymes, PCR and gel electrophoresis
• Gel electrophoresis-isolation of DNA using
  electrical currents and a gel matrix made of
  sugars

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Genetic Fingerprinting Gel Electrophoresis

• DNA has (-) charge, it will run towards the
  bottom of gel that has () charge
• Fragments separate based on size. Match fragment
  pattern with DNA found at scene.
• Take DNA found at crime scene
• Restriction enzymes, STRs, PCR
• Gel electrophoresis
• Compare DNA fragments?name suspect

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DNA Sequencing

• Allows for the actual nucleotide sequence to be
  known
• Automated, done by machines
• Used to solve all the genes in humans? Human
  Genome Project
• Is there an advantage to knowing the entire gene
  sequence of an organism?
• Functional genomics and comparative genomics

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Applications of Genetic Engineering

• In agriculture
• Strawberries that resist frost
• rice with beta carotene
• tomatoes that last longer
• pest resistant corn (95 modified)
• Larger animals? more meat, milk
• In environment
• Bacteria that clean up oil/toxic spills
• In medicine
• Insulin, growth hormone, interferon, taxol,
  relaxin, erythropoietin

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Genetic Engineering

• Problems
• Is it safe for human consumption? Allergies?
• If organisms get out into natural setting, impact
  environmental balance?
• Bigger animals?more waste, competition
• Hybrids herbicide resistance gene in corn passed
  to a weed

The Enviropig has been genetically modified in
such a manner that its urine and feces contain
almost 65 percent less phosphorus than usual.
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Gene Therapy

• Introducing the normal gene into humans with
  disease
• We can make the genes through rDNA, but how do
  we get them inside to every cell?
• Ex vivo gene therapy uses modified viruses to get
  the new gene inside cells ? SCID, familial
  hypercholesterolemia
• In vivo gene therapy uses direct injection or
  inhalation of normal/healthy gene to defective
  cells/organs
• How do we prevent an immune reaction?