Part:BBa_K2974410
C. elegans Trigger Sequence
The C. elegans trigger is an RNA sequence designed to be used in conjunction with a T7 promoter to induce GFP expression for Lambert iGEM’s C.elegans Toehold switch Biosensor. When the trigger RNA sequence is present, it binds to the complementary sequence on the toehold switch, expressing the GFP reporter gene and producing a green fluorescence. The trigger RNA sequence was designed using NUPACK (Nucleic Acid Package) software. Our trigger (36 base pairs) assembled with the T7 promoter was then inserted into pSB1A3, which is a high copy plasmid obtained from the parts registry.
Description
Toehold switches are riboregulators that activate translation in response to a distinct RNA sequence. It is comprised of a switch and a trigger. The switch is composed of a hairpin loop structure that represses translation through its complementary bases in between the ribosome binding site and start codon, which is followed by a 21 nucleotide linker sequence. These sequences ensure that the toehold switch structure will be maintained while coding for low-molecular weight amino acids that would not interfere with the switch’s function. The toehold domains at the beginning of the hairpin are 12 to 18 nucleotides long and are designed to be complementary to the trigger in order to initiate linear RNA binding. The trigger contains complementary sequences to the toehold domain that once it is in the presence of the switch, it will bind to the hairpin stem and unbind the loop. This exposes the ribosome binding site and start codon, allowing translation of the reporter protein to occur.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 25
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