Team:Cambridge/Project/FluorideRiboswitch

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Previous iGEM teams have charaterised an impressive array of inducible promoters, along with other elements of biosensing circuitry... Read More






Fluoride riboswitch

We also plan on implementing and characterising a Fluoride riboswitch. This, unlike the Magnesium construct, is a positive regulator. The riboswitch, originally isolated from Bacillus cereus, serves as a transcriptional attenuator in the abscence of fluoride. In the presence of fluoride its conformation changes and the repression is lifted. In B. cereus this serves to permit translation of a fluoride efflux pump, which allows the bacteria to cope with the potentially toxic fluoride levels in which it finds itself.

We have kindly been provided with a plasmid from the Breaker lab in Yale University, the key features of which are shown below:

Our riboswitch construct for the detection of fluoride.

This construct was initially tested with an X-GAL assay, which produced excellent results. Having gained this qualatative data, we moved onto performing a Miller assay to produce more quantatative data.

File:X-GalAssay.jpg
The results of our X-GAL assay on the fluoride riboswitch construct. Numbers on the top of the tubes represent the fluoride concentration of each solution in mM.

If time permits, we are also aiming to put our part into Ratiometrica, once it is completed:

References

  • Jenny L. Baker et al., Widespread Genetic Switches and Toxicity Resistance Proteins for Fluoride, Science 335, 233-235.

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