Designing C2H2 Zinc Finger Proteins to Target Specific Genomic Sites

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Designing C2H2 Zinc Finger Proteins to Target
Specific Genomic Sites
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• Peter Zaback
• Iowa State University
• Rocky 06

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Simplified zinc finger binding model
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Application Gene repair/insertion via induced
homologous recombination

• Two zinc finger binding domains with attached
  nuclease monomers can be designed to bind
  adjacent target site on the DNA
• This brings the nuclease monomers together to
  form an active nuclease dimer, cleaving the DNA
  at the target site
• Cleavage greatly enhances homologous
  recombination which can be used to insert/replace
  genetic material
• This allows us to insert genes, repair mutations,
  knockout existing genes

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Zinc Finger Targeter (ZiFiT)
http//bindr.gdcb.iastate.edu/ZiFiT
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Is there a simple explanation for our preliminary
results?
Segal, D.J. et al. PNAS. 1999 v. 96 p. 2758-63
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Future Work

• Initial experiment Construct fingers of varying
  composition for all 27 possible three-finger
  combinations of strong, average, and weak binding
  
• Construct additional variants to test our ability
  to predict the affinity of designed zinc finger
  arrays for their target sites

Acknowledgements

• Drena Dobbs
• Dan Voytas

• Jeff Sander
• Fengli Fu
• David Wright
• Ronnie Winfrey
• Jeff Townsend

Zinc Finger Consortium http//zincfingers.org
Funding USDA MGET, NIH, CIAG (ISU)