Reporter

Part:BBa_K494000

Designed by: Haissi Cui, Tilman Flock, Sebastian Gude, Christoph Hartlmüller, Florian Praetorius, Jan Schüürmann, Tobi Wauer, Philipp Wortmann   Group: iGEM10_TU_Munich   (2010-10-25)

Malachite Green Binding Aptamer

Methods to visualize nucleic acids via fluorescence are rare, partly due to the size of fluorescent reporters. Thus, we present the malachitegreen-binding aptamer to the parts.igem. By adding only 38 bp, fluorescent determination of specific nucleic acids becomes possible allowing evalutation of PoPS-based devices via in vitro transcription. Binding of triphenyl dye malachitegreen to the aptamer increases fluorescence by 2360-fold. This leads to an significant increase and a shift in absorbance from 618 to 630 nm. With an emission maximum at 652 nm, aptamer-bound malachitegreen fluoresces at longer wavelength than most dyes and does not interfere with those. We provide this part for efficient in vitro evaluation of PoPS-based devices in general and switches based on our concept in particular.

The malachitegreen-binding aptamer has been successfully used in screening systems being both robust and easy to produce. Aptamers provide specifities in the range of antibodies and can be evolved to target small molecules and proteins. Since malachitegreen is a membrane permeable dye, its uses are not limited to in vitro measurements. The malachite green aptamer can be used to tag and follow any RNA, including messengar and small RNAs to study questions about their metabolism and biological functions.[1] Aside from the application as a mere reporter, the malachitegreen-binding aptamer has already been utilized to build up modular sensors which can together with another RNA-binding domain sense and report small molecules like ATP for example. This new detection method seems to provide promising future applications and sensors. Since the principle of modularizing fits well into our concept of building networks, we like to provide this part to allow further engineering considering in vitro sensing systems.

Fluorescence spectrum of malachite green bound to BBa K494000. Excitation: 630 nm

Sequence and Features

Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 1
    Illegal BamHI site found at 33
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


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