'To design and characterise a toehold switch with potential to detect any given RNA sequence. Our immediate target is the detection M. bovis RNA in a safe, low-tech and cost-effective manner'
The primary aim of the Ribonostics project is to design a toehold switch (a type of riboregulator) with the potential to detect any given RNA sequence, and to standardise it into a BioBrick for future use by other iGEM teams.
Tuberculosis (TB) in cattle is a serious problem local to our region, the South West, causing devastating economic and personal losses to farmers in the dairy and beef industries. The current test for bovine tuberculosis(bTB) interferes with the BCG vaccine, giving a false positive result if the animal has been vaccinated. As a result, using the BCG vaccine in cattle is illegal in the EU. Therefore, the immediate application of our project is the detection of Mycobacterium bovis, the causative agent of tuberculosis in cattle. Our test will detect RNA secreted by M. bovis into the bloodstream, expressing a reporter if this RNA is present. As our test is RNA-based, it will not interfere with the BCG vaccine, and we hope that with time the BCG vaccine can be brought back, at least in the UK. Hence we are able to differentiate between infected and vaccinated animals.
In order to make this a simple test which can be used in the field, we are aiming to express it in a cell-free system. When designing our test we must also consider the people using it, hence we plan to integrate their thought and the current methods. We aim to use chromoproteins as indicators in the test, and therefore we have further characterised three chromoproteins aeBlue, eforRed, and amajLime to allow for easier measurement/etc. In order to do this, we have measured OD over a range of wavelengths for each part to find their peak maxima. We also constructed standard curves of chromoprotein concentration vs. OD to allow for easier quantification of chromoprotein expression, and determination of visual limits to determine the amount of protein required for the colour to be intense enough to easily observe. To see the results of this, see the experiment's page