Escherichia coli

We have used E. coli as a chassis organism to allow us to clone our gene construct successfully. After using Golden Gate cloning to make our Level 1 and Level 2 constructs, we needed to transform E. coli with the newly made constructs. This is to ensure that we have enough of the correct construct when we get to the next stage of transforming Agrobacterium tumefaciens.

Agrobacterium tumefaciens

A. Tumefaciens (Agro) was a chassis organism in this project to act as a shuttle chassis for the delivery of our Level 1 and Level 2 DNA constructs into Nicotiana benthamiana. Agro infects the plant through its Ti plasmid. This inserts a segment of its DNA, T-DNA, into the host’s genome. This allows the expression of the gene of interest.

Nicotiana benthamiana

N. benthamiana is a widely used experimental plant from the solanaceous group of flowering plants that includes tomatoes, potatoes and capsicums. It is widely used in plant pathology due to the large number of plant pathogens (viruses, bacteria, fungi, oomycetes etc) that can successfully infect it. Of importance to synthetic biologists, N. benthamiana it is easily genetically transformed and regenerated and amenable to facile methods for virus-induced gene silencing and transient protein expression including the production of therapeutic compounds and pharmaceuticals. We selected it as the plant host as it is a model organism for the plant world.