Team:Mingdao/Safety

All personnel receives a tour and are informed of the rules upon entering the lab, the main rules include:
1. No running, sudden moves within the lab.
2. No eating within the premises of the lab.
3. All experiment waste must be thrown in a special bin then put through a machine for sterilization.
4. All pipettes must be sanitized after use.
5. All equipment use must follow the using manual.
6. The mosquito cage must always be double sealed.
7. A bottle of ethanol must always be around when handling the mosquitoes.
8. To enter the P2 lab, all personnel must be thoroughly sanitized, must wear full lab suits, remove shoes and watches, and pass through two doors.

CO2 analysis:

In order to prove the seriousness , we measure the CO2 concentration in enclosed spaces and analysis the results.

If the concentration of CO2 is between :

● 350~1000 ppm, the air is fresh
● 1000~2000 ppm, people feel the air turbid and start feeling drowsy
● 2000~5000 ppm,people feel headache,lethargy, sluggishness, heart beat hasten, mild nausea
● above 5000 ppm, it may cause severe hypoxia, coma..

picture(1)

picture (1) is the result of our measure in a simple single room (only one person inside an enclosed space with air condition on .) According to the picture, the concentration had reached to 1000 ppm in a hour. While the subject felt asleep, it slowly declined(We speculate CO2 may leak out from the gaps, and the rate of venting is bigger than the rate of producing.) Until getting up next day, the speed of breathing was back to normal.

picture(2)

picture(2) is the result in a regular car with four people in. In eleven minutes, the concentration in the car had increased from 2353 ppm to 4660 ppm.

The result shows if staying in a unventilated room for a long time will have bad influences to our health. (the more people there are, the more CO2 is produced.It may increase the rate of CO2 production and make the problem even worse.) The problem of excessive CO2 concentration is indeed widely exists in our daily life and may have badly influenced our health.So it is important for people to solve these problems.

Current method:

we have gone to National Chung Hsing University, Taichung, Taiwan to search for current methods and come up with a better way.

Physics-Physical absorption:

(1) Using organic or inorganic solvents(For example: Propylene Carbonate,, and ocean water.) as absorbents
(2) Using solid absorbents, such as carbon or Zeollte to absorb carbon dioxide.
● Disadvantages:
(1) The absorption is deteriorated due to the sulfide, causing the number of regeneration to lessen.
(2) The absorption isn't effective

Chemistry-Chemical absorption:

● The chemical reaction between the absorbent and carbon dioxide is used to recycle CO2,and the reverse reaction is used to regenerate the absorbent.
● Disadvantage:There are many operational variables, making it more cumbersome to operate.

Biology-biological absorption:

● Taking advantage of photosynthesis in plants.
● Disadvantages:
(1)The efficiency is low
(2)The caring is time-consuming.
(3)The growing condition of the plants are unpredictable

Our approach:

Since the solubility of CO2 is low in water, we combine biology and chemistry method, inserting the gene that can produce carbonic anhydrase(CA, which can increase the solubility of CO2 in water) into bacillus subtilis. Then use light and high photosynthesis efficiency algae to purify the water quality.

Comparison:

Physics and chemical methods must be done in lab, while our approach can be used in any place. The physics method needs very large space in the application place especially, comparing to our device, it’s light and portable.

	Requirement	Advantage	Disadvantage
Physical absorption	(1) high pressure (2) low temperature	(1) can be used again (2) the higher the pressure is, (low temperature) the higher the efficiency is	(1) demand is difficult to meet (2) only suitable for places with high CO2 pressure
Chemical absorption	chemicals	high efficiency	(1) can’t be used repeatedly (2) by products is corrosive
Biological absorption	(1) sunlight (2) air (3) water	(1) environmental friendly (2) low risk	(1) low efficiency (2) caring is required
Our approach	electricity	(1) portable (2) easy to clean (3) increasing the solubility of CO2 in water	need to clean every day

Experiment design:

We use Bacillus to secrete enzymes out of the bacteria to achieve continuous production of CA enzyme without destroying the cell wall. As for algae, we choose cyanobacteria to match the appropriate temperature and illuminance to achieve better result.

Procedure:
1. Insert the gene that can produce carbonic anhydrase(CA) into bacillus subtilis
2. Use light and high photosynthesis efficiency algae to purify the water quality.

Gene cloning:

Part collection:

pBS0EPliaI promoter / pBS0E
BGSCID: ECE742
PliaI is an inducible promoter from B. subtilis, and it was first developed in The Bacillus BioBrick Box 2.0.We have got this part from BGSC(http://www.bgsc.org/).

shine dalgarno / pSB1C3
Sequence:AAAGGAGGAA
It’s within the primer we designed to replicate the CA (Synpcc1792_1388 & Sypcc1447) sequence.

CA (Synpcc7942_1388 & Synpcc7942_1447)
We have got the algae which has the CA sequence from National Chiao Tung University, Taiwan. With PCR(Polymerase chain reaction), the CA part is able to connect with the primer. To compare the different rate of dissolving carbon dioxide, we used Synpcc7942_1388 & Synpcc7942_1447 sequence respectively.

Terminator / pSB1C3
part: BBa_B0015
This is the most commonly used terminator in gene cloning with the consisting of Double terminator BBa_B0010 and BBa_B0012. We also get this part from iGEM Jamboree in its Spring 2019 Distribution-2019 Kit Plate 3.

Expected results:

● reduce the concentration of CO2 in enclosed space efficiently
● portable
● moderately sized
● intelligence control is available (IoT)
● algae in the device can be used again

Conclusion:

In order to solve the high concentration of carbon dioxide in enclosed space, we invented our product to deal with the problem simply (without special equipments). The followings are the methods and the advantages of our device.

Application:

Since our device is portable, it can be used everywhere.The followings are the places that often contains high CO2 concentration and can be easily improve by using our device.

● public transportation
● classroom
● gym
● office
● cafeteria
● house

Signification:

● Differ from physics or chemistry way, our product can work without special experimental equipments
● Portable and can be used everywhere
● The device operates in a circulation way and is easy to clean up.
● The algae we produce can be used in different aspects(ex.fertilizer)
● Improve indoor air quality in an environmentally friendly way

Product design:

Reference:

Chen, P.H., Liu, H.L., Chen, Y.J., Cheng, H., Lin, W.L., Yeh, C.H., Chang, C.H., 2012. Enhancing CO2 bio-mitigation by genetic engineering of cyanobacteria. Energy and Environmental Science 5, 8318–8327.