Team:Imperial College/B.subtilis

Throughout this summer we worked with B. subtilis as a chassis. Although this chassis has great potential, it is not commonly used and very few B. subtilis BioBricks are available. We aimed to increase the accessibility of B. subtilis as a chassis. The major results of our work are summarised on this page.

In order to pursue our project we had to design and build a variety of BioBrick parts for B. subtilis. These can be split up into three groups:

• Promoter and Ribosome Binding Sites (RBS) - These promoter and RBS combinations include a number of inducible and constitutive promoters combined with weak and strong ribosome binding sites.
• Coding Regions - These parts include specific coding regions concerned with our project such as biomaterials, and more general coding regions such as antibiotic resistance genes that can be used for selection markers.
• Integration Sites - As an alternative to maintaining genetic devices on extra-chromosomal plasmids we investigated the construction of parts to allow integration of genetic devices into the B. subtilis chromosome.

In order to expand the use of B. subtilis as a potential chassis we investigated the different protocols used for the transformation of B. subtilis. B. subtilis is used as the model for gram positive bacterium and have been extensively studied. A variety of protocols have been developed for the transformation of B. subtilis. We investigated two different protocols, one relying upon natural competence of stressed B. subtilis and the other using electroporation. The protocols and results were both fully documented to further the use of B. subtilis as a chassis.

• The protocols can be found here - Protocol 1 and Protocol 2
• The summary of results can be found here - Summary of results

Growth Curve
We characterised the growth rate of B. subtilis in LB media at 37^oC. This curve provides useful information when inducing protein synthesis in B. subtilis and when characterising parts such as promoters. In addition to collecting this growth curve information, we modeled the growth rate of B. subtilis which can be found in the clicking Growth Curve section of the Dry Lab.