BBa_K1586002
1
ZeusJSTOP
EsxB synthetic toehold switch J23100 STOP
2015-09-17T11:00:00Z
2015-09-18T12:22:18Z
Based on the toehold design by Green et al. 2014.
===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 attached to the toehold switch is expressed.
A toehold switch is unique compared 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 changed to detect any given trigger RNA molecule, and the protein coding region can be swapped for any desired reporter protein for easy measurement/visualisation.
Toehold switches can be used to detect specific RNA molecules in a cell-free system, or 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 K1586000 encodes for a toehold switch forward engineered by Green et al. (2014), nicknamed GreenFET1 (Green forward engineered toehold). GreenFET1 is designed to detect a synthetic RNA trigger (GGGACUGACUAUUCUGUGCAAUAGUCAGUAAAGCAGGGAUAAACGAGAUAGAUAAGAUAAGAUAG) and produces GFP when activated. It contains non-standard ribosome binding site (RBS) and GFP reporter protein and has an intended use as a control/comparison toehold. For characterisation and use, GreenFET1 has been put under the control of a J23100 promoter. It can also be found under the control of a T7 promoter as part <partinfo>K1586001</partinfo>.
===Characterisation===
false
false
_2003_
24485
24485
9
false
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
false
Exeter iGEM 2015
annotation2469617
1
GGG leader sequence
range2469617
1
34
36
annotation2469615
1
J23100
range2469615
1
1
33
annotation2473491
1
TAA
range2473491
1
114
116
annotation2469622
1
Protein coding region
range2469622
1
102
869
annotation2469620
1
Switch region
range2469620
1
37
61
annotation2469621
1
B0034
range2469621
1
87
98
annotation2469625
1
Terminator
range2469625
1
878
1006
annotation2469624
1
GFP (E0040)
range2469624
1
150
869
BBa_K1586002_sequence
1
ttgacggctagctcagtcctaggtacagtgctagcgggggaccccggcctgacgaatataagtcgtttaaaaaaaaaaatataattaaagaggagaaacgaatgatattcgtctaaagtggtagtggtagtggtgaatgcgaatgcataatgcgtaaaggagaagaacttttcactggagttgtcccaattcttgttgaattagatggtgatgttaatgggcacaaattttctgtcagtggagagggtgaaggtgatgcaacatacggaaaacttacccttaaatttatttgcactactggaaaactacctgttccatggccaacacttgtcactactttcggttatggtgttcaatgctttgcgagatacccagatcatatgaaacagcatgactttttcaagagtgccatgcccgaaggttatgtacaggaaagaactatatttttcaaagatgacgggaactacaagacacgtgctgaagtcaagtttgaaggtgatacccttgttaatagaatcgagttaaaaggtattgattttaaagaagatggaaacattcttggacacaaattggaatacaactataactcacacaatgtatacatcatggcagacaaacaaaagaatggaatcaaagttaacttcaaaattagacacaacattgaagatggaagcgttcaactagcagaccattatcaacaaaatactccaattggcgatggccctgtccttttaccagacaaccattacctgtccacacaatctgccctttcgaaagatcccaacgaaaagagagaccacatggtccttcttgagtttgtaacagctgctgggattacacatggcatggatgaactatacaaataataaatttaaatccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata
igem2sbol
1
iGEM to SBOL conversion
Conversion of the iGEM parts registry to SBOL2.1
James Alastair McLaughlin
Chris J. Myers
2017-03-06T15:00:00.000Z