Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Exeter iGEM 2015
Date created: 2015-09-17 11:00:00
Date modified: 2015-09-20 07:33:46
EsxB toehold switch J23100
| Types | DnaRegion |
| Roles | mature_transcript_region RNA |
| Sequences | BBa_K1586003_sequence (Version 1) |
Description
Usage and Biology
A toehold switch is a type of RNA molecule known as a riboregulator/riboswitch. It is able to detect the presence of a specific ssRNA molecule (termed the 'trigger RNA') which has a sequence complementary to its switch region through base pairing. If the correct RNA molecule is detected, the protein coding region of the toehold is expressed.
A toehold switch is unique in comparison to other types of riboswitches as it is completely synthetic, and therefore easier to engineer and standardise. The fact that the toehold switch can be modified means that the switch region can be easily changed to detect any given trigger RNA molecule, and the protein coding region can be swapped for any desired reporter protein most suitable for its application.
As shown by Exeter iGEM 2015, toehold switches can be used to detect specific RNA molecules in a cell-free system. In addition to this, the plasmid DNA encoding the toehold switch can be transformed into cells in order to ascertain whether a gene is being expressed (through detection of its mRNA). The applications of this technology can range from a research tool (e.g. detection of secreted RNA in cell supernatant, detection of gene expression, etc.), through to more commercial/medical applications such as diagnostic testing.
The Part
Part K1586003 encodes for an adaptable and standardised synthetic toehold switch. This toehold switch was built to contain a standard promoter (J23100), RBS (B0032), and GFP (E0040). It was designed to be used in a diagnostic test, with an initial target of the EsxB gene mRNA. EsxB encodes for Esat6 and is involved in the pathogenicity of Mycobacterium bovis. For more on this issue and the use of toehold switches in diagnostic testing, visit Exeter iGEM 2015's wiki (http://2015.igem.org/Team:Exeter).
Characterisation
Notes
The first 12 nucleotides of the toehold (not including the GGG leader sequence) should not form a part of the toehold stem.The majority of the RBS should be within the loop region of the toehold.
There must not be an inframe stop codon within the main linker sequence.
The linker sequence length must be a multiple of three.
For a more in depth explanation, see the design page of our wiki: http://2015.igem.org/Team:Exeter/Toehold_Design
Source
Synthetic sequence| Sequence Annotation | Location | Component / Role(s) |
| J23100 GGG leader sequence Switch region B0034 Protein coding region GFP (E0040) Terminator Stem-loop | 1,33 34,36 37,61 87,98 102,869 150,869 878,1006 51,113 | promoter feature/promoter feature/misc sequence_feature feature/misc sequence_feature ribosome_entry_site feature/rbs feature/cds CDS CDS feature/protein sequence_feature feature/misc stem_loop feature/stem_loop |