From 2009.igem.org

Team Request

iGEM Judges, The Alberta 2009 iGEM team has selected a large-scale cloning and assembly project, with the ultimate goal being to build a complete functional E. coli chromosome in vitro from a near-minimal set of essential native E. coli genes. In support of this goal, our team has designed a new standardized in vitro assembly method, named Biobytes. The system is based on a bead platform where constructs with long sticky ends are annealed together sequentially. This system is designed to enable the single day assembly of plasmids composed of up to about half a dozen premade parts. Complementary to existing BioBrick standards for DNA assembly, we expect this system to be useful for rapid generation of large constructs wherever an interpart spacing of 37 bp can be tolerated. A detailed description of our method is attached.

  • We are requesting a variance in order to deliver the following DNA from this project, which we believe will be sufficient for future teams to take advantage of our system:Two universal acceptor plasmids (named pAB and pBA) useful for cloning genes to assemble using this strategy
  • A number of individual E. coli genes cloned into these plasmids

8A small set of useful promoters, reporters and selectable markers cloned into the plasmids

Using these plasmids as well as universal primer sequences for the PCR extraction of bricks from the plasmids anyone will be able to use this system for their own rapid assembly. The design ensures that the existing catalog of BioBrick parts is compatible with the system, and can be easily cloned into the universal plasmids for brick generation. We believe that this new system is ready for both use and formalization, and we are working on a standard document to submit as a BBF RFC. Thank you for your time and consideration.


David Lloyd University of Alberta iGEM Team

U of A Variance Request