Composite

Part:BBa_K1132031:Experience

Designed by: iGEM Toulouse   Group: iGEM13_INSA_Toulouse   (2013-09-19)
Revision as of 01:22, 5 October 2013 by Truan (Talk | contribs)

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In vitro characterization of BBa_K1132031

The basic idea is to perform the switch with a cell lysis containing the recombinase and the plasmid containing the gate to be switched. After incubation, transformation of the plasmid containing the gate allows the quantification of switched versus non switched plasmids. Experiments have been performed as described in the protocol. http://2013.igem.org/Team:INSA_Toulouse/contenu/lab_practice/notebook/protocols/charac_recomb

Results

In the absence of recombinase, the colonies are white. As a result, AND gate does not have transcriptional leakage and non-specific recombinases cannot switch the door.

A1.PNG

We tested the AND gate with Bxb1. After incubation with Bxb1, the colonies are still white (this is normal!), therefore visual inspection of the switch was not possible. The only possibility was to sequence independent clones We analyzed five independent colonies by sequencing, all of them were switched.

689px-Switch_AND_1.png


This part can be considered as a good amelioration of the RFP reporter system. Reporter systems can be controlled either by regular positive or negative regulators. Such regulation systems obey to biological rules and are modulated between to limit values. The logic gates obey a second type of rule, closer to the electronic description of the 0 and 1 (Off and On) states. Therefore, the switch are less prone to physiological states than regulatable promoters. Furthermore, we also demonstrated that the switch are permanent and can be genetically transmitted to the offspring. This also constitutes a new system for determining at any time any occurring event that can further disappear. The switch would serve as a tracer. Last, as these switches obey Boolean logic operations, detection of independent, time separated events can also be achieved. A full description of the logic gates is present on our [http://2013.igem.org/Team:INSA_Toulouse/contenu/lab_practice/results/logic_gates Wiki].



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