Improved BioBrick Standard

For a multitude reasons we would like to work with the upgrade of 2007 Freiburg iGEM proposed standard (Assembly standard 25). The upgrade includes altered multiple-cloning site which enables friendly scar after part ligation and simple extension of linker between parts.

Advantages:

• • in-frame fusion of protein parts
  • benign protein scar/scars
  • preserving standard restriction sites of prefix and suffix
  • four new restriction sites are added in multiple-cloning site
  • heat inactivation??
  • no Dam methylation problem for XbaI
  • stand-alone protein expression
  • full BBb compatibility and
  • blunt-cutting isochizomer of NgoMIV (NaeI) and XmaI (SmaI) possibility of directional cloning with two

restriction enzymes enables part transfer between different formats and other potentially interesting transfer reactions

Disadvantages:

• • unexpected site effects for users not aware of different prefix/suffix
  • N-parts could be assembled with different enzyme combination
  • not compatible to BioFusion format (frame shift; stop codon)
  • not compatible to BBb format (Berkeley format)
  • additional limitations on the nucleotide sequence to avoid additional restriction sites

Here we describe sequence properties of modified vector BioBrick-NIC-II. All sequences defined herein are specified in the 5' to 3' direction.