Difference between revisions of "Team:OUC-China/Safety"

 
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             <div style="display:inline;height:45px;color: white;padding:0%;white-space:nowrap;text-decoration:none;position:relative">MODEL
 
             <div style="display:inline;height:45px;color: white;padding:0%;white-space:nowrap;text-decoration:none;position:relative">MODEL
 
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                <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model" style="text-decoration:none;padding:0;">Overview</a></div>
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              <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model" style="text-decoration:none;padding:0;">Overview</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#Ordinary" style="text-decoration:none;padding:0;">ODE</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#ode" style="text-decoration:none;padding:0;">ODE</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#Thermodynamic" style="text-decoration:none;padding:0;">Thermodynamic model</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#nt" style="text-decoration:none;padding:0;">Thermodynamic model</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#Riboswitch" style="text-decoration:none;padding:0;">Riboswitch</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#stablizer" style="text-decoration:none;padding:0;">Stablizer</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#sta" style="text-decoration:none;padding:0;">Stablizer</a></div>
 
                
 
                
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#asRNA" style="text-decoration:none;padding:0;">Antisense RNA</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#asrna" style="text-decoration:none;padding:0;">Antisense RNA</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#Molecular" style="text-decoration:none;padding:0;">Molecular Dynamics</a></div>
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                 <div class="nav-sub-item"><a class="kkk" href="https://2019.igem.org/Team:OUC-China/Model#md" style="text-decoration:none;padding:0;">Molecular Dynamics</a></div>
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<li class="nav-item8" style="width:180px;height:54px;padding:9px;">
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             <a style="display:inline;height:45px;color: white;padding:0%;color: white;text-decoration:none" href="https://2019.igem.org/Team:OUC-China/Collaborations">COLLABRATIONS</a>
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             <a style="display:inline;height:45px;color: white;padding:0%;color: white;text-decoration:none" href="https://2019.igem.org/Team:OUC-China/Collaborations">COLLABORATIONS</a>
  
 
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<div style="position:absolute;z-index:98;width:100%;background:white"><img src="https://static.igem.org/mediawiki/2019/2/22/T--OUC-China--description_top.png" style="max-width: 100%;max-height:100%"></div>
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<div style="position:absolute;z-index:98;width:100%;background:white"><img src="https://static.igem.org/mediawiki/2019/5/5d/T--OUC-China--Safety_top.png" style="max-width: 100%;max-height:100%"></div>
 
<div style="position:fixed;text-align:center;max-width:6%;z-index:100;top:-10px;left:46%;right:53%"><a href="https://2019.igem.org/Team:OUC-China"><img href="https://2019.igem.org/Team:OUC-China" src="https://static.igem.org/mediawiki/2019/9/9a/T--OUC-China--home_logo.png" style="width:108px;"></a></div>
 
<div style="position:fixed;text-align:center;max-width:6%;z-index:100;top:-10px;left:46%;right:53%"><a href="https://2019.igem.org/Team:OUC-China"><img href="https://2019.igem.org/Team:OUC-China" src="https://static.igem.org/mediawiki/2019/9/9a/T--OUC-China--home_logo.png" style="width:108px;"></a></div>
 
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<div class="zhiqi" style="display:inline-block;width:100%;vertical-align: top;text-align:center;">
  
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<div class='text' style="font-weight:bold;color:#9ad3c8">We are still like the Wright Brothers, putting pieces of wood and paper together.</div></br>
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<div class='text'>1. Safe project design</div></br>
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<div class='text' style="font-weight:bold;color:#9ad3c8;text-align:right">——Luis Serrano</div></br>
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<div class='text'>Being responsible for our earth and ourselves is our attitude towards every experiment. In our project, all the parts we have utilized are selected from Risk Group 1, none of which are with a Red Flag. What’s more, nonpathogenic bacteria, <i>E.coli DH5AlphaZ1</i> is employed as the main chassis. As a matter of fact, when using modular riboswitch to tune the expression of genes, it is harmless for the growth of bacteria and relatively safe for industrialized applications. For the sake of environments, all the chassis organisms will be sterilized before being abandoned to prevent the genes from leaking.</div></br>
 
<div class='text'></div></br>
 
<div class='text'></div></br>
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<div class='text'>By snapping together various pieces of different colors, shapes and sizes from a Lego box, a multitude of structures with different functions such as a boat, a car, and a building can be readily built. In the ideal world of synthetic biology, biological parts such as genes, promoters, and terminators are analogously treated as Lego blocks. </div></br>
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<div class='text'>2. Safe lab work</div></br>
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<div class='text'></div></br>
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<div class='text'>We works at a neat lab named “Lab for Microinnovation and Enterprise”, which is a BSL-2 laboratory. Before working in the lab, we are supposed to be trained, not only for the general lab safety rules, but also for the standard experimental operation avoiding unnecessary damage. </div></br>
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<div class='text'>However, the key challenges in synthetic biology in real life exist on two main levels. One is the modularization and standardization of biological parts, while the other is the integration of these biological parts into devices with desired functions. Unlike Lego blocks, many of the existing parts are still incompatible and unpredictable, whose variability will crash the system sometimes. </div></br>
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<div class='text'><img src="https://static.igem.org/mediawiki/2019/7/7c/T--OUC-China--safety1.png" style="max-width:100%"></div></br>
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<div class='text'>So biologic parts of “Lego-ization” are necessary.</div></br>
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<div class='text'>Fig.1 Our laboratory and personal safety precautions</div></br>
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<div class='text'>There is no zero risk to the experimenter in any experiment, hence we are properly equipped with facilities, such as latex gloves, nitrile gloves，goggles, lab coats to protect us from biotic and abiotic hazards.</div></br>
 +
 
 +
<div class='text'>Furthermore, several operational zones are divided scientifically for special experiments like A zone where is weighing the drugs, preparing the medium and sterilizing, B zone where is the recording experimental note and organizing experimental data area, C zone where is a molecular operation area, D zone where is RNA electrophoresis and a polyacrylamide gel electrophoresis area. For us, lab safety is a hot topic and we have come up with a practical lab safety manual which normalizes our lovely lab. </div></br>
 
<div class='text'> </div></br>
 
<div class='text'> </div></br>
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<div class='text'>Since their discovery, riboswitches have been attractive tools in bacterial systems. Natural riboswitches are found with the highest frequency in the 5’-UTR of bacterial mRNAs, they have two main components: an “aptamer domain” and an “expression platform”. in response to the binding of a specific target molecule, they can regulate the expression of downstream genes through structural changes. Also, more artificial riboswitches are engineered to regulate the expression of proteins of interest. </div></br>
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<div class='text'><img src="https://static.igem.org/mediawiki/2019/e/ef/T--OUC-China--safety3.png" style="max-width:100%"></div></br>
 +
<div class='text'>Fig.3 Safety operation</div></br>
 
<div class='text'></div></br>
 
<div class='text'></div></br>
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<div class='text'>The useful application of riboswitch</div></br>
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<div class='text'>Practical lab safety manual</div></br>
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<div class='text'></div></br>
 
<div class='text'></div></br>
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<div class='text'>•  Routine inspections must perform between 6:30-8:00pm and 9:30-11:00pm.</div></br>
 +
<div class='text'>•  Lab cleaning must be done twice a week with disinfectant moping the floor for at least 3times.</div></br>
 +
<div class='text'>•  Well prepared experiment plans should be written before you start.</div></br>
 +
<div class='text'>•  Lab coats and gloves must be worn when conducting experiments.</div></br>
 +
<div class='text'>•  Short skirts, shorts, and open shoes must not be worn.</div></br>
 +
<div class='text'>•  If you see a colleague doing something dangerous, point it out to him or her.</div></br>
 +
<div class='text'>•  Lab coats must not be worn outside laboratories and in public areas.</div></br>
 +
<div class='text'>•  Know where safety equipment (eyewash, shower and extinguisher) is located.</div></br>
 +
<div class='text'>•  Food/drink is not allowed in laboratories where chemicals are used stored.</div></br>
 
<div class='text'></div></br>
 
<div class='text'></div></br>
 +
<div class='text'>3. Safety Education</div></br>
 +
<div class='text'>OUC-China has built up a WeChat public platform, a worldwide platform with billions of users for documentation, communication and popularization. To engage with the public and help them attach importance to bio-safety,we published some articles. Fortunately, all these articles are rather approachable and most of them received positive feedbacks.</div></br>
 
<div class='text'></div></br>
 
<div class='text'></div></br>
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<div class='text'></div></br>
 
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<div class='text'></div></br>
 
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<div class='text'>But due to context-dependent performance and limited dynamic range, the use of riboswitches is often restricted. </div></br>
 
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<div class='text'><img src="https://static.igem.org/mediawiki/2019/6/62/T--OUC-China--Description2.png" style="max-width:100%"></div></br>
 
  
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<div class='text'></div></br>
 
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<div class='text'>All in all, the three problems above make the riboswitch quite hard to design and employ to the application, which make it can’t be regarded as a modular device. Towards to the three problems, now the exiting strategies are describing following by:</div></br>
 
−
<div class='text'> </div></br>
 
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<div class='text'>~ To make the riboswitch as a modular plug-and-play device, scientists try to insert a sequence between the riboswitch and gene of interest to protect the structure of riboswitch from damage so that we can change the CDS easily.  The sequence they have chosen is by random design and test by some high-throughput screening method such as SELEX. </div></br>
 
−
<div class='text'> </div></br>
 
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<div class='text'>~ To change the response functions of riboswitch, scientists try to design the expression platform by biology method and rational design. Many studies used the directed evolution  to optimization the dynamics range of specific riboswitch and develop the bio-physics model  to design it.</div></br>
 
−
<div class='text'> </div></br>
 
−
<div class='text'>~To change the concentration of ligand in the experimental culture system, scientists often use the physics-based method, such as microfluidics device  or replacing the media with fresh non-inducer-containing media .</div></br>
 
−
<div class='text'></div></br>
 
−
<div class='text'>By reviewing the exiting problem and solutions towards them, we are aware of some aspects worth optimizing. Adding a redundant sequence before the GOI directly will lead to the expression of fusion protein which may destroy the GOI's structure and function. And the method such as random design and directed evolution may waste too much time to achieve the goal you desired . While the effect of the microfluidics device is various between different labs. </div></br>
 
−
<div class='text'></div></br>
 
−
<div class='text'>This year, OUC-China proposed a standardized design principle named “RiboLego” which can break the deadlock we have mentioned before, making the riboswitch a modular, tunable one and easy to toggle between the on and off state. We hope our design will make it easier and more efficient for future igem teams to get the expected expression by using riboswitch .</div></br>
 
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<div class='text'></div></br>
 
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<div class='text'>We divide modular riboswitch into three parts: the original riboswitch, Stabilizer, Tuner from 5' to 3'. </div></br>
 
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<div class='text'style="text-align:center"><img src="https://static.igem.org/mediawiki/2019/f/f2/T--OUC-China--Rdesign2.png
 
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" style="max-width:60%"></div></br>
 
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<div class='text'>Stabilizer is a sequence which can prevent the structure of the riboswitch from damage. It has a clear source to generate and the appropriate length designed by <a class="click" href="https://2019.igem.org/Team:OUC-China/Model
 
−
">model</a>.</div></br>
 
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<div class='text'></div></br>
 
−
<div class='text'>Tuner placed between Stabilizer and the GOI to split them from each other has a function that reduces the expression probability of fusion protein and avoids destroying the GOI's structure and function. What's more, designed by <a class="click" href="https://2019.igem.org/Team:OUC-China/Model
 
−
">model</a>, Tuner can be used to control the riboswitch function precisely, achieving the desired level of expression.</div></br>
 
−
<div class='text'></div></br>
 
−
<div class='text'>We validate our design principle in different riboswitches including three kinetic switches: Adda riboswitch, Btub riboswitch, cobalamin biosensor, and one thermodynamic switch:  Four U riboswitch. What's more, three different kinds of GOI is used including sfGFP, YFP, and mRFP1. The good results show the high universality of our design principle.</div></br>
 
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<div class='text'></div></br>
 
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<div class='text'>To toggle between the on and off state of kinetic switches, we use the model to design different asRNAs which target different region to activate or deactivate the riboswitch. We will  optimize this system continuously and finally achieved to regulate the on-off state of riboswitch.</div></br>
 
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<div class='text'></div></br>
 
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<div class='text'></div></br>
 
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<div class='text'>All in all, inspired by the three existing blocks, we design our alternative riboswitch design frameworks, 'RiboLego', to make the riboswitch modular, tunable, reliable and time-saving.</div></br>
 
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<div class='text'></div></br>
 
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<div class='text'>Click <a class="click" href="https://2019.igem.org/Team:OUC-China/Medal
 
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">here</a> to get more information about our achievements!</div></br>
 
  
  

Latest revision as of 08:22, 30 November 2019

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1. Safe project design

Being responsible for our earth and ourselves is our attitude towards every experiment. In our project, all the parts we have utilized are selected from Risk Group 1, none of which are with a Red Flag. What’s more, nonpathogenic bacteria, E.coli DH5AlphaZ1 is employed as the main chassis. As a matter of fact, when using modular riboswitch to tune the expression of genes, it is harmless for the growth of bacteria and relatively safe for industrialized applications. For the sake of environments, all the chassis organisms will be sterilized before being abandoned to prevent the genes from leaking.


2. Safe lab work

We works at a neat lab named “Lab for Microinnovation and Enterprise”, which is a BSL-2 laboratory. Before working in the lab, we are supposed to be trained, not only for the general lab safety rules, but also for the standard experimental operation avoiding unnecessary damage.


Fig.1 Our laboratory and personal safety precautions

There is no zero risk to the experimenter in any experiment, hence we are properly equipped with facilities, such as latex gloves, nitrile gloves，goggles, lab coats to protect us from biotic and abiotic hazards.

Furthermore, several operational zones are divided scientifically for special experiments like A zone where is weighing the drugs, preparing the medium and sterilizing, B zone where is the recording experimental note and organizing experimental data area, C zone where is a molecular operation area, D zone where is RNA electrophoresis and a polyacrylamide gel electrophoresis area. For us, lab safety is a hot topic and we have come up with a practical lab safety manual which normalizes our lovely lab.



Fig.3 Safety operation


Practical lab safety manual


• Routine inspections must perform between 6:30-8:00pm and 9:30-11:00pm.

• Lab cleaning must be done twice a week with disinfectant moping the floor for at least 3times.

• Well prepared experiment plans should be written before you start.

• Lab coats and gloves must be worn when conducting experiments.

• Short skirts, shorts, and open shoes must not be worn.

• If you see a colleague doing something dangerous, point it out to him or her.

• Lab coats must not be worn outside laboratories and in public areas.

• Know where safety equipment (eyewash, shower and extinguisher) is located.

• Food/drink is not allowed in laboratories where chemicals are used stored.


3. Safety Education

OUC-China has built up a WeChat public platform, a worldwide platform with billions of users for documentation, communication and popularization. To engage with the public and help them attach importance to bio-safety,we published some articles. Fortunately, all these articles are rather approachable and most of them received positive feedbacks.