Prof Duncan Shaw

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Lecture 44

• Prof Duncan Shaw

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Recombinant DNA technology

• First technical breakthrough in medical genetics
  was chromosome analysis in 1950s
• Second is recombinant DNA, in the 1970s - 1990s
• This will culminate in the complete DNA sequence
  of humans (the Human Genome Project)
• There are now many methods available to analyse
  patients DNA in the lab, to identify mutations,
  to discover new genes, etc.

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How to purify a gene

• First method is by cloning, i.e. introduce the
  gene into a bacterial cell then grow up large
  amounts and extract DNA (in vivo)
• Second method is by polymerase chain reaction
  (PCR) using DNA polymerase to amplify the gene in
  a test-tube (in vitro)
• Both methods have their uses but PCR is preferred
  in medical applications because it is quicker and
  cheaper

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Bacteria provide the means

• Bacteria have been vital in developing DNA
  technology
• Thermus aquaticus (which lives in hot springs)
  provides DNA polymerase enzyme for PCR
• Escherichia coli (which lives in our guts)
  provides plasmids (mini-chromosomes) used in
  cloning
• 100s of bacterial species provide restriction
  enzymes that cut DNA at specific sequences of
  bases (4 - 8 bases long)

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Applications

• In biomedical research - to identify the genes
  responsible for human characteristics (including
  disease)
• To analyse what goes wrong with these genes in
  disease (pathology)
• To provide prenatal and presymptomatic diagnosis,
  carrier detection, risk calculation
• New therapies (drugs, gene therapy)

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95o
50o
72o
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1 2 3
1.3kb
1.1kb
Diagnosis by DNA of sickle-cell anaemia
1 normal 2 carrier (heterozygote) 3 affected
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DNA sequencing by the Dideoxy (Sanger) method
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