Device

Part:BBa_K3702192

Designed by: Dat Vo, Christine Cremeans   Group: iGEM20_FSU   (2020-09-28)


ParE2 generator

This device is designed to generate ParE2 toxin. This was used to make a Toxin/Antitoxin Kill switch. It can be paired up with a ParE2 generator to make a kill switch that would only kill the host cell when an inducer is not present/present.

We decided to model our kill switch system after team UCAS 2016 where they designed a TA module with the toxin ParE2 and antitoxin ParD2 that is dependent on the amount of tetracycline in the wastewater treatment plant, in low concentration of tetracycline, the gene will stop producing the ParD2which leaves ParE2 by itself to kill the host bacteria. We took this kill switch design and made changes that we thought would be better for our problem. First we built a design with a stronger RBS and a different terminator. The second change was switching out the promoter. Their design used a tetracycline inducible promoter Ptet. While this promoter work for them, it might not work as well for us since our project was based on the U.S wastewater treatment plants while their project was based in China, it was also risky using this promoter since we can not confirm how much the amount of tetracycline fluctuate and that it would be the same throughout different plants in the U.S. So we decided to go with something that is more likely to work, a methane inducible system. This system was more likely to work since methane is already abundant in wastewater treatment plants since they have to treat human feces, whereas a tetracycline inducible system was dependent on something that was more out of control.

For more details on the Sewage Purification Limiting Antibiotic Spread in a Habitat (SPLASH) project, visit https://2020.igem.org/Team:FSU/Engineering

Figure A: Design specification of the kill switch using SBOL Visual, ParE2 Generator with the Formaldehyde Inducible ParD2 Generator (BBa_K3702206)


Usage and Biology

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 340
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


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Categories
Parameters
None