From 2010.igem.org

Revision as of 19:06, 27 October 2010

Project overview by animation

Motivation

Our motivation arised from the following emergent needs in the development of synthetic biology:

• Detailed design rules for large-scale genetic circuit design.
• Comprehensive information of the interactions among genetic parts in vivo.
• Exploring gene expression mechanisms using traditional methods takes too much time.

The Biobrick Parts Registry is beginning to overflow with parts. At last count, there are over 14000 parts in the registry. With this many parts, the registry has made it a very easy to design experiments, but for its complexness it has made it very hard to complete experiments. We design experients, order the parts, receive the parts, and ligate them together, only to find failed results. Even if we follow the correct arrangement of parts (i.e. regulator-RBS-coding sequence-terminator), some parts still don't fit together in a circuit.

Yet, the current iGEM trend is to create larger and larger circuits, circuits that have less and less chance of working together. We've seen iGEM teams get more creative with bacteria, with more ideas that can enhance synthetic biology, but they often ends as only ideas and designs. Rather than spending time hoping that the parts we want will fit together, we need to find out how parts interact so that we know which parts work with each other.

Just like we know the design rules that tell us how IC components work with each other to create a working circuit, we need to find similar design rules in synthetic biology. The lack of these rules creates limitations in the development of synthetic biology.

Current research uses the expression of reporter genes to tell when a circuit is working.We want to quantify gene expression in both space and time, so that we can better study the interaction between different biological parts in vivo. However, studying these gene expression mechanisms using current methods takes far too much time.

With these problems in mind, we created SpeedyBac.

Overview

For iGEM2010, the NYMU-Taipei team has created a novel assaying system ("SpeedyBac") that can

• speed up the expression detection of a gene flow.
• reveal the location and quantity of both mRNAs and Proteins.
  • Between the mRNA level and protein level of our gene expression cycle, we have integrated a riboswitch that allows us to stop, start and control the translation of proteins. Using this switch, we can study mRNA and its protein(s) in one cycle without the interference of one on the other.
• the speedy degradation device we built can stop the gene expression quickly and cleanly.

Design

To achieve our goal, our SpeedyBac system is designed with the following three devices:

• Speedy switch
  • Controls and speeds up mRNA translation into protein via a riboswitch between mRNA and protein level of gene expression.
• Speedy reporter
  • Using mRNA aptamers and split GFP-eIF4A reporter systems to quickly outperform promoter-only activity.
• Speedy protein degrader
  • Fast, specific, and constitutive proteolysis achieved by engineering fluorescent proteins with LVA tags.

Safety Issues

Here we detail how we approached possible issues of biological safety associated with our project.

Specifically, the following four questions were considered:

1. Would any of our project ideas raise safety issues in terms of:
   • researcher safety,
   • public safety, or
   • environmental safety?
2. Is there a local biosafety group, committee, or review board at our institution?
3. What does our local biosafety group think about our project?
4. Do any of the new BioBrick parts that we made this year raise any safety issues?
   • If yes, did we document these issues in the Registry?

Our answers to these four questions:

1. For iGEM 2010 project, our goal is to design and engineer bacteria (called "SpeedyBac") to provide a faster assay system for exploring the design rules of synthetic biology. However, due to potential safety issues of using pathogenic bacteria in our experiments, we only used Escherichia coli model organism in our experiments to prove the concept. Although these viral proteins we used are viral capsid proteins for binding to huamn cellular receptors, they are neither toxic nor pathogenic by themselves. These viral capsid proteins or even viral paticles are not able to replicate in the bacterial chassis we used for making ViroCatcher. Therefore, they should not raise safety issues in terms of:
   • researcher safety,
   • public safety, or
   • environmental safety.
2. At NYMU, we do have a biosafety committee to review all biosafety and biosecurity issues at our university.
3. We had presented our ViroCatcher project to many of our school professors including many of the members of our biosafety committee. Since we were not using viral particles or viral vectors in any of our experiments, they did not think the use of viral capsid proteins in our project would raise any biosafety issues.
4. None of the new BioBrick parts that we made this year raise any safety issues. Since we are not authorized to give out those gene clones of viral capsid proteins, all those clones are not shipped as new BioBrick parts at this time.

We also documented all our answers to these safety questions in our presentation, wiki presentation, and poster.

Acknowledgements

We are grateful for the kind support and help of

• [http://www.southampton.ac.uk/biosci/about/staff/cgp1x07.page Dr. Chris Proud] for providing us with the pGEX-eIF4A of his lab for our experimental use. Dr. Christopher Proud is currently a Professor of Cellular Regulation & Deputy Head of School, Research School of Biological Sciences Life Sciences Building University of Southampton Southampton, UK.
• We thank the campus faculty and students for their suggestions and for their comments on this iGEM project.
• National Yang Ming University and Ministry of Education, Taiwan. This iGEM project is fully supported by them. We wish to acknowledge and thank their supports of this project.