When we develop our strains, in order to help more people with depression solve the problems they have with diseases, we are as commercialized as possible and can be manufactured in factories on a large scale. As a result of the above philosophy of this project, the strains we designed have been affected from engineering design to future expansion marketing.We have strategies in place to target users, improve the user experience, secure projects, and how to make projects more accessible in the real world.

  Since the team designed a synthetic biological strain based on the production of 5-HTP and GABA based on E. coli Nissle1917, this strain can be very good at reducing fear, anxiety and stress, good sleep quality, lowering blood pressure, improving patient mood, improving sleep quality, and relieving pain. As health care products for the general public, probiotics should not only be limited to the fixed depressed patients, but should be widely available to the general public. The strains developed by our team can be used not only for patients suffering from mild to moderate depression, epigliasis, loss of appetite, easy sighing, insomnia and many dreams, but also for people who live in a high-pressure and fast-paced lifestyle for a long time, office workers and students, etc.

  The drug adopts the three-layer coating technology of cyclodextrin, which is an oligosaccharide composed of amylose and will not be decomposed by proteases in the stomach. The presence of hydroxyl group on the outside makes it hydrophilic, and the C-H bond on the inside of the cavity makes it hydrophobic. The above properties can allow drug particles to dissolve well in water, and relatively intact from the digestive tract to the intestine for lysis. In order to further improve the survival rate and efficacy of probiotics, we summarized the following points for attention:

Use warm water less than 40 degrees Celsius to dissolve the drug particles

  The optimal survival temperature of our probiotics is 37 degrees Celsius, which is to adapt to the temperature in the human gut, so that it can better adapt and survive in it. But if the drug is breasted at too high a temperature, it is likely to kill the probiotics. Even though the cyclodextrin coating technology can separate the inside from the outside, it does not prevent heat transfer.

Use pure water for brewing

  Pure water is the ideal solution for drugs. Milk, carbonated drinks, tea, coffee and other drinks have different properties, and the pH value and contents of different drinks may affect the solubility and integrity of drugs. In particular, tea brewing solution was avoided because we also inserted protocatechuic acid-specific promoters linking lytic genes into probiotics. Once catechuic acid was detected by probiotics in the gut, the bacteria would automatically dissolve to release 5-HTP and GABA, but at the same time, a large number of bacteria would die.

The amount of water for each packet should be 200ml

  200ml water is the most suitable, because too little water will cause adverse results such as drug caking, which will affect the solubility and absorption of drug particles in water. If there is clumping, you can add a certain amount of water to fully dissolve and enhance the activity of probiotics.

Not recommended for diabetics

  The coating substance used is mainly cyclodextrin, which is a sugar, so diabetics need to pay attention to their sugar intake.

  In case of sudden sharp increase in pressure, insomnia and emotional excitement, you can brew a cup of tea and take it to relieve symptoms quickly. This is because drinking tea is a good traditional culture and can be well accepted by the public. At the same time, it can make people relax, and the caffeine contained in it can refresh themselves. Moreover, the tea polyphenols will be decomposed into protocatechuic acid (PCA) after entering the intestine. We set the corresponding protocatechuic acid promoter in probiotics to connect the bacterial autolysis program. Tea polyphenols and phenolic acids in tea will be metabolized to protocatechuic acid (PCA). We used protocatechuic acid (PCA) -specific promoters to ligate lytic genes. We selected the SRRz lysing gene to create our lysing module, which consists of S, R, and Rz. After consumers take our probiotics, the probiotics produce GABA, 5-HTP in the gut. When needed, we take tea leaves, and the protocatechuic acid in tea leaves starts the lysis system. After bacterial lysis, GABA and 5-HTP will be released in the intestine, which can achieve the purpose of alleviating the symptoms of depression in a short time.

  Microbial factories are widely used to produce a wide variety of chemicals, food, drugs and energy, and are the core link of biological manufacturing. With the improvement of biotechnology, many people transform microbial cells into microbial factories that can produce materials from low-value renewable resources by using diverse biochemical reaction networks of microorganisms and re-engineering metabolic pathways. So far, MCFs has been able to produce antibiotics, amino acids, recombinant proteins and so on, which are widely used in pharmaceutical, food, energy and agriculture fields. For our project, in the construction of cell factories, we have transformed in vitro chemical synthesis into E. coli Nissle 1917. For the synthetic pathway, we also screened the plasmid gene sequence, inserted the required target gene, and constructed the complete cleavage system and suicide system by using the promoter, operon and other gene elements, which can efficiently and safely complete the assembly of functional modules and the creation of synthetic pathways. For the optimization of cell performance, because Escherichia coli is mainly anaerobic environment, NISSLE1917 can also show good activity in the small intestine, but in order to improve the stability of gene links, we need to provide good temperature, pH and other superior conditions for cell transformation and reproduction. At the same time, in order to ensure the high yield of bacteria in vivo, we need to simulate the temperature, pH value and other conditions in the small intestine in vitro to optimize the cell quality and better achieve the purpose. That way, we could get the similar cell factories that produce artemisinin, C3 and so on, to produce the probiotics we need. But conventional cell factories are mass-produced in vitro. Our design is to maximize the reproduction of the modified NISscle1917 in vitro, and then transplant the process produced in the factory into the body.

  Further investigation of Nissle1917 revealed the existence of Nissle 1917 not only in humans but also in animals. According to the data, we found that Nissle1917 can also play a role in the animal body, and has great advantages. Porcine Nissle1917 can stabilize the intestinal tract for a long time and play a biological function of immune regulation. We propose to repeat the experiment we did in human Nissle 1917 on animal Nissle 1917, in which the human tryptophan hydroxylase I (TPH1) gene and glutamic acid decarboxylase gene are transferred to express 5-HTP and Gaba, and complete the design of gene suicide system and encapsulation. To make it work in animals as well as humans. In this way, the mental state of animals taking psychoprobiotics will be greatly improved, so as to improve their meat output quality or milk yield, etc., which has a good development for the current world's animal husbandry.

Avoid genetic contamination

  For the control of the suicide system, our plan was to engineer a genetic suicide system that would allow E. coli Nissle 1917 to kill itself after leaving the small intestine. This section has been tested and discussed extensively in the past. Here's a new idea: we could look for a probiotic that's only found in the gut (it dies when it leaves the body, it's never been found in the wild) and engineer it so that it produces two neurotransmitters, 5-HTP and GABA, to do its job. Since it can no longer live in the wild, there is no need to worry about genetic contamination.

Coating technology

  The coating technology of probiotic strains can ensure that the activity of probiotics is not destroyed to the maximum extent, so that the probiotics can resist gastric acid and choline smoothly into the intestine. After entering the intestine, the probiotics can be released under the specific environment in the intestine, so that a large number of live bacteria can successfully colonize the intestine and function. We used the unique multi-layer probiotic coating technology to protect the probiotics, so that the probiotics can play a greater role in human health. It includes a probiotic membrane, a lipid membrane, and a polymer membrane to protect E. coli.，which is resistant to heat, saliva, bile, and other digestive fluids. Based on this theory, we have designed special wraps that can give better treatment to patients taking this probiotic. In conclusion, there is no safety problem with the probiotics we designed and produced.

Efficiency

  There are still many questions about the idea of putting probiotics into animal farms, such as how efficiently the animal-derived Nissle1917 can produce two substances, whether the antitoxin gene and toxin gene can be matched in the design of the gene-suicide system, and how well they are expressed, which requires experimental manipulation and exploration, which is an avenue to explore in the future.

Bias

  Nowadays, many people are prejudiced against genetically modified products. For them, genetically modified products, which cannot be proved to be absolutely safe, are dangerous. Most of the public's fear of GMO comes from fake articles or untrustworthy wechat official accounts, many of which are groundless and illogical but highly inflammatory. As a result, more and more people are afraid to buy the products we envision. Therefore, we should further strengthen the education of the masses, let all people slowly accept and use our products, and let more patients with depression get help and relief.