Genetic Techniques and Drosophila melanogaster

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Genetic Techniques and Drosophila melanogaster

• Brian Stephenson
• Rachel Pileggi

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Why the Fly?

• 61 of human disease genes have homologs in the
  fly
• Genes have only one copy
• The amount of crossing over is minimal and only
  occurs in females
• Approx. 13,600 genes in Drosophila versus
  40,000-60,000 in humans
• Genes are more similar to mammals than nematodes
• Half of fly proteins show homology with mammalian
  proteins

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Balancer Chromosomes

• Multiple overlapping inversions
• Markers and lethal recessives
• Allows for all heterozygous in stock

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P-Element Transposons

• TransposonsDNA that can move to different
  positions within a single cell
• P-Element ends are used with cDNA to create
  vectors
• Activated by transposase enzyme

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P-Element Transposons
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Hybrid Dysgenesis

• P-elements with hybrid dysgenesis serve as
  transformation vectors and as genetic tags
• Vectors use P-element ends and chosen cloned DNA
  to insert into eggs of mother
• Genetic tags make vector and mutations easy to
  follow

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Identifying Genes with Enhancers

• Enhancer Trapping introduce genes within
  P-Element Transposons
• E. coli lacZ gene is placed downstream of
  promoter
• lacZ codes for B-Galactosidase, reporter
• P-Element plasmid only expressed with tissue
  specific enhancer

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Identifying Genes with Enhancers
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Genetic Mosaics to Determine Gene Function

• FLP site-specific recombination enzyme from
  yeast
• Introduced (using P-elements) downstream of
  heat-shock promoter whose expression is turned on
  at elevated temperatures
• Synthesizes FLP recombinase which catalyzes
  recombination
• Result high frequency of clones new mutations
  identified by effects in homozygous patches

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Genetic Mosaics to Determine Gene Function
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Ectopic Expression

• Expression outside the cells or tissue where the
  gene is normally expressed
• Introduced into germ line
• Can be induced with heat shock protein or
  mechanism with GAL4 and UASg cDNA as with the
  lacZ gene

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Ectopic Expression