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This year, the ZJU-China iGEM team aims to design and realize a tunable RNA scaffold to accelerate biosynthesis pathways and turn their on and off. As one of the most vital biomacromolecules, RNA plays a crucial role not only in coding process, but also in non-coding one. RNA scaffold is designed to colocalize enzymes through interactions between binding domains on the scaffold and target peptides fused to each enzyme in engineered biosynthesis pathways in vivo, which may suffered from low efficiency of production caused by relative lack of spatial organization of non-homologous enzymes. The scaffold allows efficient channeling of substrates to products over several enzymatic steps by limiting the diffusion of intermediates thus providing a bright future for solving the problem. Meanwhile, we plan to add an aptamer structure on RNA scaffold as a riboswitch to regulate biosynthesis pathways by micromolecular ligands. Then we can control the all-or-none binding relationship between the enzymes and RNA scaffold by whether the special ligands are presented or not.
Safety
a. Researcher safety
We have not worked with any materials that raise risk to researchers' safety or health. We only worked with level 1 safety organisms and they are E.coli K12 stains BL21DE3, DH10β and DH5α.
Since our researchers used to face regular bio-safety challenges in a molecular microbiology lab, such as staining DNA agarose gel with EB, our lab chooses gel red as nucleic acid dye this year, which raise little if not no safety challenge. There is also a container for us to pour culture fluid and culture medium full of cells in, and thus it will be treated properly afterwards.
Any researchers conducting experiment were instructed about the right method both by printed protocols and in the lab. After the training, we passed an on-line test about the lab safety, standard experiment procedures and lab waste treatment. During any experiments, necessary individual protections such as gloves and white robes were strictly required. Also, the public safety of the lab is guaranteed by the strict registering protocols on the operation of any apparatus. In order to make sure all the requirements of safety were carried out well by the researchers and to get prepared for any emergency, our lab (also is the open bio-lab of the college) set up 24hr*7 camera system in the rooms.
b. Public safety
As we said above, what we used are non-pathogenic E. coli strains. We engineered the cells with plasmids, which contain resistance genes of antibiotics, so chances are that the genes may transmit to other pathogenic bacteria and cause some risk to public safety. To avoid it happening, we gather all the culture fluid and culture medium with cells for unified treatment and not let cells get out of our lab through sewer.
c. Environmental safety
We care about the environmental safety. We make sure no media containing engineered cells go into the sewer before sterilization, and all utensil contacted with the engineered bacteria will be sterilized. The acid and basic chemicals are neutralized and pour into the sewer. Toxic and hazardous chemicals are collected and treated properly according to the safety rules of our university.
d. BioBrick safety
The gene products of some biobricks are RNAs and they will not be translated into proteins. As we know, RNA can be easily degraded, so our biobricks will not raise potentially safety problems.
The two other biobricks express the subunits of GFP, and these products are not bio-hazard, and the expression is under effective control.
e. Biosafety provisions
Our experiment is designed and carried out following the standards of national biosafety office, especially the <Safety Administration Regulation on Genetic Engineering> (Only Chinese version is available online), as well as the laboratory regulations of our university, the college of life sciences, and the biology lab center.
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