Part:BBa_K2582000

miniSpinach (P1-a5-b3)

This Biobrick contains the DNA sequence of the light-up RNA Aptamer, miniSpinach.

Usage and Biology

miniSpinach is a minimal version of the Spinach aptamer developed by Paige et. al. (2011). It is created previously by generating mutants of the Spinach aptamer by decreasing stem lengths (Ong et. al. 2017). When transcribed in vitro, it allows the docking of fluorogen 3,5-difluoro-4-hydroxybenzylidene imidazolinone (DFHBI) and increases its quantum yields by hundreds of folds.

Characterization

miniSpinach can be in vitro transcribed or expressed in E. coli.

Mixing the Spinach aptamer with excess DFHBI yielded a 100-fold increase in the fluorescence level of the reaction mixture. However, its fluorescence in E. coli is hard to visualize and quite close to the E. coli autofluorescence level. Upon comparison between In-cell, total RNA and in vitro synthesized RNA, it is found that miniSpinach does not fold well in E. coli.

Fluorescence of miniSpinach under Chemidoc SYBR Green Mode

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]