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Nitric Oxide Sensing & The Hybrid Promoters | The Comparator Circuit | Theoretical Projects

SUMMARY OF IT ALL HERE; similar to front page

. Produced new promoters - 2 BioBricks

. Ligated these to fluorescent proteins - 4 more BioBricks

. Promoter+FP BioBricks were characterised using growth studies and fluorometer, flow cytometry and FACS

. Aim to characterise further, such as finding maximum level of induction by certain compounds and difference in induction based on chemical used.

. Future applications in cancer and environment.

Contents 1 Origins of the Idea 2 Preliminary Lab 3 Lab Work 4 Results 5 Conclusions 6 Future Applications 6.1 Cancer 6.1.1 Warrior Cell 6.2 Buildings 6.3 Soil 7 Future Experiments

Origins of the Idea

. UEA uses NO

. Difficult to sense (machines and biologically)

. Important applications e.g. cancer and buildings so useful to work it out

Preliminary Lab

. Looked at PyeaR and CArG, ligated them together in two orientations for a flexible promoter (info on what each is)

Lab Work

. Made into biobricks (link to registry page)

. Ligated to RFP/CFP and got photos to prove it works

. Used RFP/CFP with the fluorometer etc. to get data on sensitivity

. Tranfected into mammalian cells to show flexibility

. Studied the growth rate

Results

. Graphs from flow and fluorometer

. Pictures etc. from mammalian transfection

. Growth study on MB/BM

Conclusions

BM/MB better or the same, RFP/CFP better or the same etc.

Future Applications

Cancer

Warrior Cell

A problem with current cancer therapeutic techniques is that they are not specific to cancer cells, leading to patient discomfort. We hope to engineer E.coli cells that selectively identifies cancerous cells through their anaerobic environment and secrete high concentrations of NO, mimicking macrophages, and thus killing these cells. These cells are known to be hypoxic, and hypoxia is a hallmark for cancer cells already used in tumour detection. Our Warrior Cell construct hopes to pave the way for a new kind of therapeutic strategy that delivers a therapeutic agent only when and where it is needed.

Buildings

Soil

Future Experiments

. Full quantitative analysis to see where the values lie; combine with tuners for different sensitivity levels

. With different substrates (e.g. nitrite salt, NO donor)

. With different reporter/effector enzyme