This year, we have tried our best to turn our project from an idealistic concept to implementation. However, we believe that there is still a long way to go to improve our project. We have listed several strategies for further project improvement. We expect our project can make a significant impact on the world in the future.

We cloned bsNOS into E. coli in order to make it produce nitric oxide. We chose bsNOS as a target gene because Bacillus subtilis is the most closely-related species that can produce a substantial amount of Nitric Oxide. Eventually, we managed to engineer E. coli to produce Nitric Oxide. In the future, we would like to try cloning nitric oxide synthase from GH3 pituitary cells^[1]. The NOS from GH3 has a lower K_m value with higher efficiency. It may increase the nitric oxide production efficiency of our engineered E. coli.

In the current design, we used CI857, the heat-sensing protein as a switch that regulates the promoter pR. Therefore, we need to culture our engineered bacteria at 30°C to keep the switch off. This led to low culture efficiency. According to research, mutant pR enable the culture to grow under temperature up to 36°C^[2].

A mutation in the O_R2 operator region of the rightward pR promoter increases the temperature stability of the pR/cI857 gene expression system. Thus gene expression is stringently repressed at temperatures up to 36°C. By doing so, we can get better production efficiency.

Eye kNOw boasts a dynamic drug release system, one that can be applied in other situations. If we replace NOS with other enzymes, we can also develop additional dynamic treatments for intraocular pressure diseases.

Not only that, we can also add a biosensing system into our contact lenses, allowing for the detection of pathogens or inflammatory biomarkers in the eye. This way, our dynamic-drug-release system can be used for more applications. Thus, more people can experience the benefit of Eye kNOw.

We have introduced several plasmids into our engineered bacteria so it can produce not only NO but also increase its binding affinity to the contact lens. We have also established a growth switch and kill switch so our bacteria can safely function in the contact lenses. The whole system looks perfect; however, we have introduced too many plasmids into the bacteria, which might gives too much burden to the bacteria and cause harmful effects on them. Therefore, we hope that some genes can be integrated into the chromosome, while some plasmids can be merged to reduce the number of plasmids in the future.

We plan to integrate csgD, csgA, and NOS into the chromosome and merge the three plasmids of the growth switch into one, that is, clone both hicA and hicB into pCP20. This way, our total number of plasmids will be significantly reduced to one, making our engineered bacteria expression more efficient.

We hope to promote the device to the general public in the future. With Eye Screen, users can monitor their IOP level by themselves, quick and easy, wherever they are. These IOP values around-the-day will automatically be synchronized to the smartphone app, Eye Cloud, where the data will be stored and analyzed, enabling users to be informed of their risks of glaucoma in a real-time manner. It can also be placed in medical centers to increase the public's willingness to track their IOP level. Healthcare practitioners can also bring Eye Screen to long-term care institutions for IOP monitoring. We designed a compact function generator to drive the transducers, omitting the need for a conventional setup. We will also improve the Arduino signal processing settings to optimize the precision of IOP readings.

Besides, collaboration with various institutions related to glaucoma will increase the willingness to measure IOP for anyone and effectively raise awareness of glaucoma. Furthermore, while artificial intelligence (AI) in healthcare and medicine is booming, we’ll try to keep up with the trend. We hope Eye Screen will also able yo function as additional tool to collect data for machine learning. In the future, Eye Screen 2.0 will be launched with the ability to carry out big data analysis and offer better precision. This portable signal processing system could be applied in various fields, bringing benefits to society.

Eye kNOw project will be developed into a start-up company, named Eyesaac Biotechnologies. Eyesaac Biotechnologies will aim to expand the portfolio of medical products which employs our core biosensing technology to other forms of disease or medical conditions. Eyesaac Biotechnologies aims to expand the area served to cover most of the countries around the world, starting with China and Japan, which represent the largest glaucoma prevalence rates in Asia. Further expansion will be focused on the EMEA and the Americas.

The short-term goal for Project Eye kNOw is to successfully complete clinical trials and have the patents granted as soon as possible.We also aspire to change the public’s perspective on GMO synthetic biology and genetic engineering, providing accessibility and convenience to patients who require glaucoma prevention.

The long-term goals for Eyesaac Biotechnologies are to expand Eye kNOw into larger markets as well as adapting the technologies and frameworks established into treating other types of diseases such as diabetes and sepsis.