Module 4: Buoyancy

Description | Design | Construction | Characterisation | Modelling | Results | Conclusions

Design

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Gas vesicles are found largely in cyanobacteria and halophillicarchea, where they are necessary to access oxygen and light. Their formation is driven by a number of genes required for production and regulation. We are using the gas vesicle gene to attribute buoyancy to our cells so that they float on to the surface of the ocean. Our particular cluster -known as GVP is extracted from Bacillus Megaterium which contains 14 putative gens gvp-A,-P,-Q,-B,-R,-N,-F,-G,-L,-S,-K,-J,-T and -U of which the last 11 genes, in a 5.7-kb gene cluster are the maximum required for gas vesicle synthesis and function in E.coli.

Requirements:

1. The sinking of the cell should trigger the expression of the buoyancy gene.

For the purposes of our project we are using a constitutive promoter for the expression of the gas vesicle (BBa_I750016). However, our plan is to have 2 promoters one is the original starvation promoter from the partsregistry and in addition we are planning to have a heat shock promoter. This will optimise the position of the cell in the water column. As the cell sinks there will be a significant reduction of nutrients and decrease in temperature consequently this change in environmental conditions will induce the promoter and subsequently the gas vesicle formation.

2. The size and the strength of the gas vesicle must be enough to keep the cell afloat. The size and strength of the vesicle are predetermined genetically and varies from species to species. Also it is very important to say that there is a trade-off between the size of the vesicle and its strength. The smaller the vesicle the less it is prone to collapsing and vice-a-versa.

The genes we are missing in our construct are gvp-A,-P and -Q and it is worth saying that the gvp-A gene is coding for a very hydrophobic 70-amino acid long proteing that is responsible for stregthen the vesicle so the lack of this proteinmay affect the strength of our vesicle negatively. Another concern about this biobrick is its size - 5.7 kb by removing some of 11 open reading frames that may play a regulatory role.There are only 2 proteins that are known to be responsible for the structure of the gas vesicle. However some of the other open reading frames may dertermine the protein folding and the Biobrick must be carefully analysed to specify those open reading frames that are the minimum necessary for the successful formation of the gas vesicle.