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Latest revision as of 02:13, 27 October 2007

• Home
  • Site Map
  • Meet the Team
  • Fun with iGEM
  • IcGEMs@OpenWetware
  • Acknowledgements
• Infector Detector
  • Introduction
  • Specification
  • Design
  • Modelling
  • Implementation
  • Testing/Validation
  • F2620 Comparison
  • Conclusion
• Cell by Date
  • Introduction
  • Specification
  • Design
  • Modelling
  • Implementation
  • Testing/Validation
  • Conclusion
• Cell-Free
  • What is Cell-Free?
  • Cell-Free Vs. Cell
  • Our Contributions
  • Characterisation
  • Applications
• Wet Lab
  • Lab Notebook
  • Protocols & Results
  • DNA Constructs
• Dry Lab
  • Modelling
  • Data Analysis
  • Software

Using the Home-made S30 E. Coli cell extract

Aims

Proper experimental amounts for reaction mixtures of the home-made S30 E.coli cell extract.

Equipment

• Eppendorf Tubes
• Gilson p20,p200,p1000

Reagents

• Pyruvate kinase
• rNTPs
• S30 cell extract (home made)
• Reaction buffer (home made)
• Commercial S30 cell extract
• Commercial Pre-incubation mix
• Amino Acids
  • Minus Cysteine
  • Minus Leucine
• DNA from midiprep/maxiprep

Protocol

1. One reaction mixture comprises:
   • Home made S30 - 16.2ul
   • Reaction Buffer- 30ul
   • rNTP's - 1ul
   • Pyruvate Kinase - 3.1ul
   • DNA - 4ul
   • ddH₂ - 5.7ul
2. Then add 4ug/2ug worth of DNA, before topping it up to a total volume of 60ul with nuclease free water.