Team:UIUC Illinois/Project/Future

From 2014.igem.org

Future Directions

"Biology has at least 50 more interesting years. "

James Watson, 1986

Environmental Remediation

Waste Matters

Knowing that caffeine could be degraded, we wondered what became of the coffee plants that were used and abused for the world's most socially acceptable drug. Coffee production is no light matter. Large amounts of water go into coffee processing, and subsequently a large amount of waste is developed. "Commercial coffee is obtained from coffee cherries, 6% of which generate the coffee powder whereas the remaining 94% constitute the by- products such as husk, pulp water etc." [3] The massive waste production is not the only problem. It's easy to think the dry mass could be repurposed as a carbon & nitrogen source for animals and plants. However, reactions to stimulants such as caffeine are far reaching. It has been shown that caffeine can "inhibit seed germination and growth of seedlings"(Friedman and Waller 1983) as well as stunt the growth of cattle. By decaffeinating the waste, it can subsequently be used as feed and fertilizer and therefore have positive economical effects. Who knows, if coffee waste decreases and the coffee producers have an extra source of income, your next cup of coffee might be cheaper!

To ameliorate this issue of waste, we looked to different pathways that could prove utile degrading xanthine waste. One organism, Pseudomonas putida CBB1 contained a set of genes that encoded for a caffeine dehydrogenase pathway. Instead of the N-Demethylation characteristic of our aforementioned puppy probiotic operon, we explored the caffeine dehydrogenase mechanism. In contrast to N-Demethylation, this pathway is a direct method to go directly to Trimethyl Uric Acid, another "safe" compound for the host animal who cannot degrade caffeine! We began to develop primers that would isolate the cdhA, B, and C genes, and moreso amplify out the entire operon, however our PCRs were unsuccessful.

Yogurt

Let's get cultured.

The final piece to our project aimed at propagating the demethylation proteins in some sort of food vector. We figured making yogurt via Lactobacillus fermentation was our best shot. It is widely known that yogurt is made through fermentation. Species such as Lactobacillus plantarum in yogurt break down lactose and produce lactic acid, the product which gives yogurt it's texture as well as taste. Having the culture that creates the yogurt simultaneously produce assistive proteins seemed like a clear shot to us!

This is the prospective construct of our CDH plasmid. With CP25, a near constitutive Lactobacillus promoter, we aim to produce the dehydrogenase gene excessively to degrade caffeine in polluted environments.