Part:BBa_K2419003

TagCFP for analysis of elastomeric tissue

TagCFP is a monomeric protein emitting bright cyan light. It is based on the natural fluorescence of the jellyfish Aequorea macrodactyla, which expresses a GFP-like fluorescent protein. This part can be used to analyze the quality i.e. of an elastomeric tissue out of proteins connected via Catcher-Tag systems (see BBa_K2419000). While such a tissue is formed, TagCFP as a fluorescent protein can be added due to its strong fluorescent emission. As a result, the maxima of emission and absorption of the fluorescent protein shift to a higher level when excited by the specific absorption light wavelength. This shift can only be ascribed to the assembly of TagCFP to the tissue. The behavior of the emission and absorption can therefore be used as a screen of the tissue around the fluorescent protein. The tissue network can be characterized regarding its structure and density. These parameters will give information about the quality of the polymeric network.

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]

Testing the indicational function of TagCFP in an elastomeric tissue

To test the indicational function of TagCFP we created a hydrogel resembling the elastomeric tissue. We then added TagCFP both into water and the hydrogel. The results of this test are shown in Figure 1. On the right, TagCFP fluoresces in a cyan color when added to water and excited with light. Whereas the fluorescence color changes to a light blue when excited with light while being located in the hydrogel.

Figure 1: TagCFP in water (right) and TagCFP in hydrogel (left).

These results visibly show the effect of TagCFP being added to different solutions. The change of the color from cyan in water to light blue in the hydrogel indicates a shift of the emission maximum and therefore TagCFP can be used as a qualitatively test for elastomeric tissue, while additionally, further characterization of the tissue can be achieved.

Characterization by the Franconia 2017 iGEM Team

We further characterized the TagCFP to supply a broader understanding of the properties of this fluorescent protein. We therefore measured the absorption and emission of TagCFP. The regarding graph is given below (Figure 2).

Figure 2: Fluorescence spectrum of TagCFP showing absorption and emission at variant wavelengths.

The results show a difference in the wavelength of the absorption and the emission. The absorption of TagCFP has its maximum at a wavelength of 457.4 nm and the emission maximum lays at a wavelength of 478.1 nm. The overlap of absorption and emission can be seen in the area around 470 nm.

We further plotted the measured values of the excitation wavelength and the emission wavelength of TagCFP to obtain a height profile given in the diagram below (Figure 3).

Figure 3: Excitation wavelength and emission wavelength of TagCFP.

The diagram displays the distribution of the different wavelengths. The red colored area reveals the overlap of excitation and emission at its uppermost distinctness.