Team:British Columbia/Future

Table of Contents

• Further Characterization and Optimization
• Connecting with Bielefeld’s iGEM team 2015
• Future Experiments

• PROJECT

  • 
  • Inspiration
  • Description
  • Approach
  • Hardware
  • Results
  • Parts
  • Future
• LABORATORY
  • 
  • Experimental Design
  • Protocols
  • Notebook
  • Safety
• MODEL
• HUMAN PRACTICES

  • 
  • Integrated Human Practices
  • Education/Outreach
  • Collaborations
• TEAM

  • 
  • Team
  • Attributions
  • Sponsors
• MEDALS

Future Directions

Characterization and Optimization

Now that our team has found a biosensor, there are many directions our project can be taken that need to be further explored. Unfortunately, due to the time constraints of iGEM our team was not able to further test and develop our biosensor, however the next steps of our discovery are clear. Currently, our plasmids are being sent in for sequencing and when we obtain our results these sequences will be analyzed and compared to other known sequences. After this, our team plans to continue efficacy tests of our biosensor to more fully characterize our biosensor. This would include testing over longer time periods and over greater STX concentrations. Also, it is important to test the specificity of our biosensor to ensure that STX is the only compound the biosensor responds to.

After a thorough characterization, our team plans to try to optimize the biosensor to further increase its effectiveness. Much of the results we are looking for will be dependant on the results of our characterization and the weaknesses of our biosensor. This optimization process will primarily consist of optimization of our vector such as using different rbs sites and experimenting with different GFP variants. Mutagenesis is also a potential option to further optimize our biosensor.

One of the most intriguing components of our project is that we have developed a screening method that is not specific to just STX, our process and use of SIGEX can be applied to any marine biotoxins using almost identical methods. This leaves the doors wide open for what other biosensors can be discovered.

Our biosensor would then be ready to be implemented in a field setting. There are many directions this component of the project could have been taken. Primarily, our team looked into the use of cell free systems in conjunction with a team-built fluorometer in order to achieve inexpensive testing with an emphasis on the ability to create a hardware system capable of field use in remote communities.

CFPS & iGEM Bielefeld

Traditional biosensors are faced with many problems in their implementation in a real world setting as they are often have slow response times, need trained professionals to operate, require expensive and tedious storage and often can only function when paired with delicate and stationary lab equipment. In recent years cell free protein synthesis (CFPS) has been used to implement the same functionality as whole cell biosensors while drastically reducing cost and increasing response times, portability and accessibility of biosensors for the untrained public.

CFPS is a strong and feasible way our project could be implemented to meet our goal of providing safe and reliable testing of saxitoxin for rural communities. There are many ways CFPS can be accomplished. One method is through the use of a commercially available system such as NEB’s PURExpress system which is highly consistent and reliable however, one of our team’s goals was to create a low-cost biosensor so a homemade solution would be ideal.

Other iGEM teams have successfully completed and implemented this (most notably Bielefeld 2015) and have demonstrated outcomes that would be able to be directly applied to our project.

Other iGEM teams have successfully completed and implemented this (most notably Bielefeld 2015) and have demonstrated outcomes that would be able to be directly applied to our project.

CFPS is one of the most promising future directions for our project. It has all the necessary required components to be taken outside of the lab and into the real world and its feasibility has been demonstrated extensively within the literature and the iGEM community. With more time and resources this could allow our project to be successfully implemented into rural communities around the coast of British Columbia.

Future Lab Experiments

Run more efficacy tests (replicates)

• Scaling up tests
• Larger range of stx concentrations
• Longer times
• Specificity of biosensor

Confirm and start optimizing biosensor

• Using different rbs, change out the gfp
• Mutagenesis