Team:Paris Bettencourt/Experiments/SinOp
From 2011.igem.org
![](https://static.igem.org/mediawiki/2011/1/19/Tubeornottube.png)
SinOp system experiments
Abstract
Results for the SinOp system:
- We successfully BioBricked KinA (BBa_K606046) into PSB1C3, improving preexisting biobrick BBa_K144010 constructs and sent them to the registry
- We showed that our receptor is a good reporter for sporulation and characterized it.
- We were able to express kinA and trigger sporulation in rich medium in exponential phase.
Design overview
![](https://static.igem.org/mediawiki/2011/1/18/Sinop_principle.jpg)
SinOp system
More information on the design here.
Parts and biobrick system construction
Here is the cloning we made for this system:
![](https://static.igem.org/mediawiki/2011/8/83/1028_Cloning_plans_kinA.png)
We suceeded in recovering the KinA gene from the non biobricked plasmid synthetized de novo by the 2009 Newcastle team, and cloned it into a standard biobrick plasmid, pSB1C3. Then we cloned this gene in front of the pVeg-SpovG (K143051) promoter + RBS. These two constructs had bees sended to the registry into pSB1C3.
This construct has been cloned right away into a replicative plasmid for subtilis and transformed, and we are caracterizing it at the moment.
Characterization of the receiver strain from Veening
![](https://static.igem.org/mediawiki/2011/1/15/Zdjzpojdapjdz.jpg)
GFP gene is under SpoIIA promoter. SpoIIA gene is express in sporulation process downstream of the SinOperon. Under starvation conditions, cells sporulate and then express the gfp.
![](https://static.igem.org/mediawiki/2011/9/9b/Qdzqd.png)
We also measured fluorescence kinetics by spectrophotometry (TecanĀ®).
Characterizing the KinA emitter
![](https://static.igem.org/mediawiki/2011/7/79/Dqs.jpg)
We overexpressed kinA, in SB168, that triggered sporulation in exponential phase at 37 degrees. Under microscope, we can observe many of cells harboring small black spots, which are characteristic of sporulation.