Part:BBa_K2974310
C. elegans Toehold Sequence
This C. elegans toehold was built using BBa_K2550202 as the stem-loop domain for the toehold, and the target sequence domain is derived from the C. elegans lin-4 gene pre-mRNA primary transcript F59G1.6 (non-toxic). The described structure was optimized in design using the parameters described by Pardee et al. and validation using the toehold designer software developed by To et al. The design of this switch involved consideration of numerous components including free energy of RBS-Linker, MFE of various structures, illegal stop codons, etc. in order to optimize the secondary structure sequence. The corresponding sequence was then processed using the NUPACK (Nucleic Acid Package) to determine whether the appropriate secondary structure formed and whether the RBS was sufficiently enclosed in the upper loop. Following analysis of the sequence and structure, this part was experimentally tested.
Description Toehold switches are RNA-based riboregulators functioning as switches that can activate/inhibit reporter expression using a synthetic construct. In the presence of a reverse complementary RNA trigger sequence, reporter expression can be activated through the binding of the trigger sequence to the toehold switch, resulting in the uncoupling of the secondary structure, allowing the RBS to be freed from the "closed" position, allowing for translation of the reporter protein. Toehold switches are advantageous as arbitrary sequences can be utilized to regulate expression of a reporter, serving as unique tools for incorporation as biosensors.
Results
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 175
Illegal NotI site found at 6
Illegal NotI site found at 460 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 96
| None |