Difference between revisions of "Part:BBa K3431006"

Revision as of 08:19, 25 October 2020

oz21_A Toehold Switch for miR-21 Detection

Description

Thistoehold switch has been designed to open up its hairpin loop structure upon binding with miRNA-21, resulting in the translation of downstream reporter protein. The design of toehold switch can be separated into the following 5 regions from its 5' end: trigger binding sites, stem region, loop region with RBS, complimentary stem region with start codon, and linker amino acids. In our constructions of toehold switches for miRNA-21, we optimised the loop region with RBS and linker amino acids based on three articles: the original work on toehold switch (Green, A.A. et al., 2014), the adaptation of toehold switch to detect zika virus (Pardee, K. et al., 2016), and novel toehold switch design for detection of miRNA in mammalian cells (Wang, S. et al., 2019) . The loop structure from the article of Pardee, K. et al. has been truncated to 12 base pairs compared to the work from Green, A.A. et al. in order to reduce the leakage of output expression. We chose to test the 3 different loop structures and 2 different linker structures (Pardee, K. et al. and Wang, S. et al.) from the above-mentioned studies.

For this particular toehold switch (oz21_A), we incorporate the loop structure from Green, A.A. et al. and the linker structure from Pardee, K. et al..

Model

NUPACK ANALYSIS

VIENNA RNA PACKAGE

Link to our model page: https://2020.igem.org/Team:CSMU_Taiwan/Model

Experiment result

References

Green, A. A., Silver, P. A., Collins, J. J., & Yin, P. (2014). Toehold switches: de-novo-designed regulators of gene expression. Cell, 159(4), 925-939. Pardee, K., Green, A. A., Takahashi, M. K., Braff, D., Lambert, G., Lee, J. W., ... & Daringer, N. M. (2016). Rapid, low-cost detection of Zika virus using programmable biomolecular components. Cell, 165(5), 1255-1266. Wang, S., Emery, N. J., & Liu, A. P. (2019). A novel synthetic toehold switch for microRNA detection in mammalian cells. ACS synthetic biology, 8(5), 1079-1088.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

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 ===Description===
-This toehold switch has been designed to open up its hairpin loop structure upon binding with miRNA-21, resulting in the translation of downstream reporter protein. The design of toehold switch can be separated into the following 5 regions from its 5' end: trigger binding sites, stem region, loop region with RBS, complimentary stem region with start codon, and linker amino acids. In our constructions of toehold switches for miRNA-21, we optimised the loop region with RBS and linker amino acids based on three articles: the original work on toehold switch (Green, A.A. et al., 2014), the adaptation of toehold switch to detect zika virus (Pardee, K. et al., 2016), and novel toehold switch design for detection of miRNA in mammalian cells (Wang, S. et al., 2019) . The loop structure from the article of Pardee, K. et al. has been truncated to 12 base pairs compared to the work from Green, A.A. et al. in order to reduce the leakage of output expression. We chose to test the 3 different loop structures and 2 different linker structures (Pardee, K. et al. and Wang, S. et al.) from the above-mentioned studies.
+Thistoehold switch has been designed to open up its hairpin loop structure upon binding with miRNA-21, resulting in the translation of downstream reporter protein. The design of toehold switch can be separated into the following 5 regions from its 5' end: trigger binding sites, stem region, loop region with RBS, complimentary stem region with start codon, and linker amino acids. In our constructions of toehold switches for miRNA-21, we optimised the loop region with RBS and linker amino acids based on three articles: the original work on toehold switch (Green, A.A. et al., 2014), the adaptation of toehold switch to detect zika virus (Pardee, K. et al., 2016), and novel toehold switch design for detection of miRNA in mammalian cells (Wang, S. et al., 2019) . The loop structure from the article of Pardee, K. et al. has been truncated to 12 base pairs compared to the work from Green, A.A. et al. in order to reduce the leakage of output expression. We chose to test the 3 different loop structures and 2 different linker structures (Pardee, K. et al. and Wang, S. et al.) from the above-mentioned studies.
 For this particular toehold switch (oz21_A), we incorporate the loop structure from Green, A.A. et al. and the linker structure from Pardee, K. et al..