xTo build more complex cellular systems, new tools and techniques are required. We are generating standardized parts, tools, and techniques for the gram-positive chassis B. subtillis. Easy to handle and transform, this bacterium offers many adantages to E. coli', including the ability to secrete proteins and integrate DNA into the chromosome. We have designed, built, and submitted gram-positive RBSes, promoters, and shuttle vectors.As a part of this work we have confirmed single copy chromosomal insertion, demonstrated InFusion assembly, and characterized an improved GFP variant. You can find our Bacillus protocols here.
We created a codon-optimized and biobricked version of the 'Superfolder' GFP engineered by Pédelacq et al (2006). This GFP variant folds faster and glows brighter than other GFP variants. Read more...
Creating new Biobrick-compatible B.subtilis vectors
Vectors submitted: We are using the In-Fusion™ PCR method from ClonTech to create new Biobrick-compatible integrative and episomal vectors. These vectors will allow us to insert Biobricks into the Bacillus subtilis genome at the AmyE locus and as a 3-5 copy plasmid that will replicate autonomously in Bacillus.
Successful transformation of Bacillus
Transformation with episomal vectors
ECE166 : High-level constitutive expression of a Green Fluorescent Protein (with the promoter Pupp) Read more...
Transformation with integration vectors
ECE153 and ECE112 : Transformation in IA751 : Amylase test
B. subtilis Promoters Designed
Information about the B. subtilis vectors can be found here.