Creating new Biobrick-compatible B.subtilis vectors

We are using the InFusion 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.

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Testing B.subtilis promoters and RBSs

We are seeking to test the expression strength and response to inducers in several promoters in B.subtilis, in combination with different Bacillus-specific ribosome binding sites.

B. subtilis Promoter Testing with Beta-galactosidase Assay

We are aiming to characterize four different Bacillus subtilis promoters using the beta-galactosidase assay. (Protocol for beta-galactosidase assay as described here

B. subtilis Promoters to be Tested:

Pupp
Ppac
Pspac
Pxyl

Links

Chris French's protocol on transforming B. subtilis
We have last year's protocol; not sure if it's trustworthy, need to compare with online resources
Protocol from BCSG catalog
- used in Cornell bacteriology course, probably should start here

B. Subtilis Transformation Protcol - Electroporation

Bongers et al. … of a Subtilin-Regulated Expression System in Bacillus subtilis: Strict Control of Gene Expression …. Applied and Environmental Microbiology (2005)