Team:British Columbia/Protocols/GibsonAssembly


British Columbia -
UBC iGEM 2012 protocols

Gibson Assembly Protocol

This protocol closely follows the suggested protocol in the NEB Gibson Assembly guidebook.

  1. Using purified DNA, add 50-100ng plasmid and 2-3X molar concentration of the inserts, or 5X molar concentration if using inserts of 200 bp or less. Add dH2O up to 10 uL. Set up a positive control using 10uL Gibson Assembly Positive Control.
  2. Add 10 uL Gibson Assembly Master Mix to each product.
  3. Incubate at 50°C for 1 hour.
  4. Using 2uL product, transform into competent cells. Since the Gibson Assembly Master Mix contains PEG, avoid using PEG sensitive competent cells. The provided positive control results in an amp-resistant plasmid.