From 2010.igem.org

Revision as of 14:39, 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. However, every team that has used information from parts registry knows how complex it is. Even if we follow the correct arrangement of parts (i.e. regulator-RBS-coding sequence-terminator), some parts just won’t work together in a large circuit design. Current iGEM teams are favor to do a big project which has a complex circuit design. However, these excellent projects almost turn out to be just design withnot much successful experimental results ultimately. So much time is wasted trying to guess blindly at which parts/devices can interact together, and which parts/devices cannot. There are definitely needs of detailed rules for large-scale circuit design and for the future development of synthetic biology.

With this in mind, we are interested in more specific design rules of a genetic circuit. We want to look closely at the central dogma, and more specifically, at the mRNA level.

Traditionally, we find out if the circuits we made are working or not by looking at the expression of reporter genes, but it would be a lot better if we could have quantitative descriptions of gene expression in both space and time. Base on the detailed information of gene expression, we can know about the interactions between biological parts in vivo. However, studying the gene expression mechanism using traditional methods takes too much time. To reduce the needed time of dealing with this kind of problem we have come up with our project: 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 or 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.

Acknowledgements

We are grateful for the kind 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.