iGEM Paris-Saclay
Project description
Recent observations lead to the idea that genes, not in the same operon but spatially close, are highly co-transcribed, even in the absence of regulatory factors at their promoter regions.
The iGEM Paris-Saclay project aims to study the effects of DNA topology on gene expression in E.coli by answering to this question: Does bringing a strong promoter closer to a weak promoter influence the expression level of genes located downstream?
We have designed a new tool based on CRISPR/Cas9 system to bring two specific DNA regions closer. This system is composed of two different dCas9 fused with each part of FRB / FKBP12 dimerization system. Each dCas9 will target a specific DNA sequence, one on the chromosome and one on a plasmid, whereas dimerization system will promote the joining of the two dCas9 when rapalog is added. In order to assess whether or not this system works, we have also designed a new tool to visualize the interaction between both dCas9. This tool is composed of a split GFP attached to two dCas9. These two small GFP tags will interact with the complementary GFP detector only if the two dCas9 are close enough to interact.
If we obtain a higher expression level of the weak promoter with our two tools, it could lead to several useful applications. For example, we would be able to use this tool to enhance gene expression of any endogenous genes due to CRISPR/Cas9 specificity. Indeed, it would be possible to design specific sgRNA but the user should be aware about off-target activity of the CRISPR/Cas9 system.
Project Overview
Recent observations lead to the idea that genes, not in the same operon but spatially close, are highly co-transcribed, even in the absence of regulatory factors at their promoter regions.
The iGEM Paris-Saclay project aims to study the effects of DNA topology on gene expression in E.coli by answering to this question: Does bringing a strong promoter closer to a weak promoter influence the expression level of genes located downstream?
We have designed a new tool based on CRISPR/Cas9 system to bring two specific DNA regions closer. This system is composed of two different dCas9 fused with each part of FRB / FKBP12 dimerization system. Each dCas9 will target a specific DNA sequence, one on the chromosome and one on a plasmid, whereas dimerization system will promote the joining of the two dCas9 when rapalog is added.
In order to assess whether or not this system works, we have also designed a new tool to visualize the interaction between both dCas9. This tool is composed of a split GFP attached to two dCas9. These two small GFP tags will interact with the complementary GFP detector only if the two dCas9 are closed enough to interact.
If we obtain a highest expression level of the weak promoter with our two tools, it could lead to several useful applications. For example, we would be able to use this tool to enhance gene expression of any endogenous genes due to CRISPR/Cas9 specificity. Indeed, it would be possible to design specific sgRNA but user should be aware about off-target activity of the CRISPR/Cas9 system.
Welcome to iGEM 2016!
Your team has been approved and you are ready to start the iGEM season!
Before you start:
Please read the following pages:
Styling your wiki
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Wiki template information
We have created these wiki template pages to help you get started and to help you think about how your team will be evaluated. You can find a list of all the pages tied to awards here at the Pages for awards link. You must edit these pages to be evaluated for medals and awards, but ultimately the design, layout, style and all other elements of your team wiki is up to you!
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Tips
This wiki will be your team’s first interaction with the rest of the world, so here are a few tips to help you get started:
- State your accomplishments! Tell people what you have achieved from the start.
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- Start documenting your project as early as possible; don’t leave anything to the last minute before the Wiki Freeze. For a complete list of deadlines visit the iGEM 2016 calendar
- Have lots of fun!
Inspiration
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