Design
To execute our idea in real life scenario we first decided to construct a plasmid that can facilitate an extracellular expression of our protein of interest i.e. Alkyl sulfatase. We decided for extracellular expression as it provides several advantages over cytosolic production in terms of purity of recombinant protein obtained as a final product and less protease activity in the culture leading to more yield. Note, that we first tried building a device that would act as a template to which our protein of interest could be cloned.
It is difficult to accomplish efficient secretion of protein without trying a different combination of secretion tags and host-vector system. Thus, it was necessary that we choose different parameters for extracellular expression for sufficient production of our protein.