Supplementary Materials

Supplementary Materials for:

A Versatile Toolkit to Produce Sensitive FRET Biosensors to Visualize Signaling in Time and Space

Rafael D. Fritz, Michel Letzelter, Andreas Reimann, Katrin Martin, Ludovico Fusco, Laila Ritsma, Bas Ponsioen, Erika Fluri, Stefan Schulte-Merker, Jacco van Rheenen, Olivier Pertz*

*Corresponding author. E-mail: olivier.pertz@unibas.ch

This PDF file includes:

  • Note S1. Generation and characterization of the mTFP1 cp mutants.
  • Note S2. Comparison of RhoA2G with the RhoA1G and Raichu-RhoA biosensors.
  • Fig. S1. Identification and characterization of mTFP1 cp mutants.
  • Fig. S2. Schematics and further characterization of the RhoA biosensor library.
  • Fig. S3. Sequence of RhoA2G biosensor.
  • Fig. S4. Schematics and further characterization of the ERK biosensor library.
  • Fig. S5. Characterization of filters compatible with mTFP1- and Venus-based biosensors and imaging screening assay for EKAR2G biosensors.
  • Fig. S6. Comparison of RhoA1G, Raichu-RhoA, and RhoA2G biosensors in N1E-115 cells.
  • Fig. S7. Characterization of EKAR2G by epifluorescence ratio and two-photon FLIM.
  • Fig. S8. Sequences of EKAR2G design 1 and 2 biosensors.
  • Fig. S9. Cloning strategy to construct design 3 and 4 EKAR2G biosensors.
  • Legends for movies S1 to S7

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Technical Details

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Other Supplementary Material for this manuscript includes the following:

  • Movie S1 (.mov format). RhoA activation dynamics during membrane protrusion and retraction in MEFs.
  • Movie S2 (.mov format). RhoA activation patterns in leading-edge filopodia of MEFs.
  • Movie S3 (.mov format). RhoA activation dynamics in response to PDGF.
  • Movie S4 (.mov format). RhoA activation dynamics during N1E-115 neuroblastoma neurite growth and collapse.
  • Movie S5 (.mov format). RhoA activation dynamics in a growing and subsequently collapsing growth cone in an N1E-115 cell.
  • Movie S6 (.mov format). ERK activity dynamics in response to EGF stimulation in HEK293T cells stably expressing EKAR2G1 (high magnification).
  • Movie S7 (.mov format). ERK activity dynamics in response to EGF stimulation in HEK293T cells stably expressing EKAR2G1 (low magnification).

[Download Movies S1 to S7]


Citation: R. D. Fritz, M. Letzelter, A. Reimann, K. Martin, L. Fusco, L. Ritsma, B. Ponsioen, E. Fluri, S. Schulte-Merker, J. van Rheenen, O. Pertz, A Versatile Toolkit to Produce Sensitive FRET Biosensors to Visualize Signaling in Time and Space. Sci. Signal. 6, rs12 (2013).

© 2013 American Association for the Advancement of Science