Part:BBa_K5109023:Experience

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Applications of BBa_K5109023

After successfully cloning Dehalogenase II inside pJUMP29-1A ∆NdeI (BBa_K5109060), before testing our part on PFAS molecules, we performed a control enzymatic test. Dehalogenase II is known to successfully degrade fluoroacetic and chloroacetic acid, therefore we decided to set some tests that would use chloroacetic acid in order to test the correct functioning of the protein.

We tested both wild type TOP10 F' as a control and TOP10 F' cells containing our expression vector. A part of the cells containing the vector were grown in addiction with IPTG 0.5 mM to induce protein expression, meanwhile another part was grown without IPTG as a control.

All the cells were then incubated with LB medium + chloroacetic acid, and monitored for 24h.

After 24h hours, the concentration of chloroacetic acid in the medium decreased for 98% in cells that were induced with IPTG, while chloroacetic acid was degraded of 90% in cells without IPTG. Wild type cells did not show any particular change in chloroacetate concentration over time.

The results lead us to consider part K5109023 a successful surface display system for this protein. Further investigation is needed in order to confirm the correct functioning of the protein, and to subsequently test it on PFAS molecules.

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