Part:BBa_K3380153
Broccoli fluorescent RNA aptamer
Broccoli is a fluorescent RNA aptamer comprising a short ribonucleotide sequence that exhibits fluorescence upon binding to a specific fluorophore.
Characterization
The Broccoli aptamer was designed and described by Filonov et al. (2014).
The Broccoli secondary structure folding prediction as simulated by [http://www.nupack.org// NUPACK] is shown below.
Usage and Biology
It is usually used in live-cell imaging, however it can also be used in vitro. The Broccoli aptamer can be used for visualisation by gene fusion to the target of interest. Its fluorescent colour is green. Some of the most commonly used fluorophores are DFHBI ((Z)-4-(3,5-Difluoro-4-hydroxybenzylidene)-1,2-dimethyl-1H-imidazol-5(4H)-one) and DFHBI-1T ((Z)-4-(3,5-difluoro-4-hydroxybenzylidene)-2-methyl-1-(2,2,2-trifluoroethyl)-1H-imidazol-5(4H)-one). The emission spectra depends on the bound fluorophore. The DFHBI fluorophore has an excitation wavelength of 492 (nm) and an emission wavelength of 516 (nm).
Its short length allows it to be synthesized rather than being cloned using plasmids. To increase its stability and increase its folding the iSpinach can be transcribed using tRNA scaffolds such as F30 (BBa_K3380101 and BBa_K3380102). The tRNA scaffolds increases its fluorescence.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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