Security is the basis
Safety in design of our project
This year, we demonstrate a modulization design--Predator Pro, to realize mediating targeted degradation of protein directly. Inspired by our project in 2018 and 2019, we further improve the construct of the Predator, a Trim21 based proteasome dependent system, alike Trim-Away manner. We modularize the Predator into different modules and demonstrate the replaceability of Bifunctional Module through replacing the antigen-antibody interaction with other interactions, including DocS-Coh2 interaction which occurs naturally, FRP-FKBP interaction which can be mediated by rapamycin and etc., from which we realize mediating the degradation by exogenous signals such as small molecule and light stimulation. Putting safety first, we considered the safety issues as follows:
1. E.coli was used to performing molecular cloning. We choose genetically modified E.coli, which has not acquired any characteristics that enable them to produce hazardous substance or invade the immune system/other systems of human or animals, as our chassis organism in our project. They are harmless from both a personal and public healthy point of view.
2. 293T cells were used to demonstrate the feasibility of the newly constructed plasmid and the Predator Pro. 293T cells, which derive from 293cells, are transfected with gene E1A of adenovirus. Though adenovirus could lead diseases to people, the 293T cells are completely safe.
3. All genes cloned into pcDNA3.1 are non-conjugative, preventing horizontal transfer of our parts. We use CMV as promoter, which guarantees that our bacteria won’t produce large amounts of protein if not induce the expression.
4. All our parts we used are guaranteed to be safe for human and won’t cause any biological pollution to the environment.
Safety in Lab Work
1. All our operations comply with the safety regulations of laboratory safety manual.
2. Before we started the experiments, our advisors gave us a wonderful lesson about Lab safety, and the original members provided detailed and comprehensive guidance to new members to avoid mistakes in the operation. Here is the emphasis on the specification:
1) Wear white coats, masks and rubber gloves.
2) Get familiar with the functions of each equipment in the lab and can use them properly.
3) Standardize experimental operation and avoid wrong operation.
4) Have a discussion on the procedures of the experiment we will perform to get familiar with the operation steps and clarify all the details.
5) Make the equipment and the appliance prepared before we begin our experiment to avoid delaying the process of the experiment or searching for laboratory appliance when operating.
6) Reasonable and scientific treatment of experimental waste is needed to prevent pollution.
7) Calm down and get ready to deal with possible accidents when operating.
8) Separate members into small groups and groups take responsibility for different experiment steps to raise efficiency.