Mutalik, V. K. et al. Precise and reliable gene expression via standard transcription and translation initiation elements. Nat. Methods 10, 354–360 (2013).

In this publication, Mutalik and colleagues characterize the BioFab libraries of constitutive promoters and bi-cistronic designs (BCDs). The latter are ways of insulating ribosome binding sites (RBS) from the adjacent genetic context. Inserting the RBSs into an upstream coding sequence, the sequence context is standardized and translation of the upstream coding sequence allows ribosomes to unwind any secondary structure.

All leading peptides used are the same, so only one insulator is represented. The RBSs described here are also the only ones present in the repository, as RBS strength can be dependant on the coding sequence used. It is to be noted, however, that the extra RBSs in the construct will compete for ribosomes and lead to the translation of small peptides. Utilizing the same leading peptide for multiple BCDs in the same circuit is also expected to lead to evolutionary instability due to recombination.