Team:SSTi-SZGD/Hardware
SSTi-SZGD
Figure 1 :
As shown in the picture, this system consists of several parts: A water tank (1), a number of pipes (2), an air pump (3), a reaction bucket/globe (4), two storage tanks (5,6) and numbers of spray nozzles (7). [more detail show in our video]
This system has two modes, a common watering mode is for day-to-day crop watering function, and an enzyme degradation mode that is for spraying enzyme specifically
Common mode: turn on the switch in the tank (1), water flows through the pipe (2) until reaches water storage tank (5). Watering takes place through either automatic setting or manual. Water comes out from the nozzles (7) and does a normal plant watering job.
Enzyme degradation mode: when it comes to pesticide residue degradation, pipe leads to water tank (5) is turned off. Whole cell product is added to the transparent bucket with a propeller with paddles inside to facilitate cell growth. The bucket is placed in the right direction inside the insulated globe (4) (which provides both dark environment and insulated conditions for cell growth as well as enzyme production) so that water pipe can be connected at the bottom of the globe to allow liquid coming in or out. Water tank (1) is turned on to add water into the bucket, and cells are allowed to grow and express proteins for 14-20 hours (normally takes place overnight). When it is ready to spray, air pump (3) is turned on to extract media containing enzymes to flow through the pipe until reaches enzyme solution tank (6). Then whole bacterial cells will be sprayed. At this stage, enzyme solution could be at rather higher concentration than needed so water is added to dilute the solution to a suitable concentration for direct spraying.
In order to eliminate the contact the GMOs with natural environment, we plan to use gram-positive bacteria, such as bacillus subtilis, as a chassis for recombinant protein secretion in the next step of the study. Gram-positive bacteria do not possess an additional outer membrane, and cytoplasmic membrane is surrounded by a cell wall. Proteins of up to 25-50 kDa can diffuse freely through the cell wall. Once the proteins are exported across the cytoplasmic membrane, they can be released directly into the culture medium(28). Bacillus subtilis also has its own twin-arginine translocation system allowing the extracellular secretion of OPH or MHE. Extracellular secretion of hydrolase is advantageous over intracellular production, as it allows the enzymes to freely bind the extracellular substrates. In this case, to avoid spraying of live bacteria, a 0.45um filter will be added to the end of the pipe to trap bacteria cells. This 0.45 um filter should be changed every time to ensure smooth spraying. In addition, to further avoid biosafety issues, we may also consider apply cell-free based spraying system.
Pesticide residual degradation usually takes place right before harvest or pick-ups. Sprayed enzyme solution could perform degradation task directly on the crop surface, soil surface as well as dripping deeper into the soil. When it is completed, simply take out the bucket with remaining cell pellets and exposed to direct sunlight, which would activate our integrated cell suicide system by allowing supernova expressed proteins to generate reactive oxygen species (ROS) and promote cell death. After a few hours, cells should be all dead and then it is safe to dispose as ordinary rubbish. This precaution prevents potential leakage of GMOs into natural environment.
