Biotechnology -- Chap. 16.

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Biotechnology -- Chap. 16.
The use of biological systems for the production
of materials (most work is in the field of
Genetic Engineering)
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Genetic Engineering

• the act of manipulating an organism's genes,
  resulting in the alteration of cellular function
• introduce specific foreign pieces of DNA into
  host cells (often bacteria or viruses) which then
  infect host cells that eventually replicate
• cells from an organism can be extracted, injected
  with a vector that inserts its genetic material
  into the nucleus of the organisms cell, and then
  placed back inside the organism
• therefore, the cells containing the new DNA can
  be grown in any quantity and so is the protein
  transcribed by that DNA.
• less commonly, the vectors can also be injected
  directly into the body

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The Process (overview) in Bacteria

• Need to convince the host cell to accept the
  foreign DNA
• this is done by attaching the foreign DNA to a
  carrier DNA molecule called a vector.
• vectors are often bacterias plasmid
• Plasmids are molecules of DNA that are found in
  bacteria separate from the bacterial chromosome.
  They
• are small, carrying one or a few genes
• are circular
• self-replicating
• this tiny but mighty plasmid molecule is the
  basis of recombinant DNA technology.

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The Process (detailed) in Bacteria
Recombinant DNA video
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• The vector and foreign DNA are cleaved by the
  same restriction enzyme in order for the sticky
  ends of both to match and then mixed together
• restriction enzymes (also called restriction
  endonucleases) are enzymes that cut the
  phosphate-backbones of DNA at specific base
  sequences (normally 4-6 bases) called restriction
  sites

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• exist naturally in bacteria in order to cut apart
  invading viral DNA
• cut parts of DNA are called restriction fragments
  
• naming-- ex) EcoRI
• 1st letter is for genus - (Escherichia)
• 2nd letter(s) is for species - (coli)
• 3rd letter is for the strain of the organism (R)
• Roman numerals at the end typically indicates the
  order of discovery (I)

Restriction Endonucleases
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• then, the enzyme ligase is added to join the
  vector and foreign DNA together making the
  foreign DNA an integral part of the plasmid
• this new plasmid is then taken up by other
  bacteria cells by a process called transformation
• identification Often the foreign DNA that was
  just inserted has other segments of DNA that
  identify it, such as resistance to antibiotics
  and the ability to produce color.

Steps in Cloning a Gene
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Adenoviruses

• human pathogens responsible for some cases of the
  human "cold".
• modified versions of two strains are currently
  being used as vectors in gene therapy trials.
• unlike retroviruses, they do not integrate into
  the host genome and the DNA molecule is left free
  in the nucleus of the host cell
• the instructions in this extra DNA molecule are
  transcribed just like any other gene
• these extra genes are not replicated when the
  cell is about to undergo cell division

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In clinical trials, gene vectors have been used
to cure lab animals of hemophilia A and B.
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DNA Sequencing

• the determination of the precise sequence of
  nucleotides in a sample of DNA.
• dideoxynucleotides are synthetic nucleotides (4
  types, ddATP, ddGTP, ddCTP, ddTTP) that lack the
  -OH at the 3' carbon atom and are each labeled
  with a "tag" that fluoresces a different color

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• chain elongation proceeds normally until by
  chance, DNA polymerase inserts a
  dideoxynucleotide instead of the normal
  deoxynucleotide
• Each of the four dideoxynucleotides fluoresces a
  different color when illuminated by a laser beam
  and an automatic scanner provides a printout of
  the sequence

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