Biosensor expression plasmids

Our previously published plasmids are available from Addgene, a non-profit plasmid repository:

Plasmids for bacterial expression

roGFP2-His in pQE-60

Grx1-roGFP2-His in pQE-60 [1]

roGFP2-Orp1-His in pQE-60 [2]

Plasmids for yeast expression

cyto-roGFP2-yGrx1 in p415TEF [4]

cyto-roGFP2-yGrx2 in p415TEF [4]

cyto-roGFP1-iL-hGrx1 in p415TEF [4]

cyto-roGFP2-Tsa2ΔCR in p415TEF [6]

cyto-roGFP2-Tsa2ΔCPΔCR in p415TEF [6]

Plasmids for insect expression

cyto-Grx1-roGFP2 in pUAST [3]

mito-roGFP2-Grx1 in pUAST [3][5]

cyto-roGFP2-Orp1 in pUAST [3]

mito-roGFP2-Orp1 in pUAST 3]

cyto-Grx1-roGFP2 in pCaSpeR4:alphaTub84B[3]

mito-roGFP2-Grx1 in pCaSpeR4:alphaTub84B [3][5]

cyto-roGFP2-Orp1 in pCaSpeR4:alphaTub84B [3]

mito-roGFP2-Orp1 in pCaSpeR4:alphaTub84B [5]

Plasmids for mammalian expression

cyto-Grx1-roGFP2 in pLPCX (retroviral) [1]

cyto-roGFP2-Orp1 in pLPCX (retroviral) [2]

mito-Grx1-roGFP2 in pLPCX (retroviral) [1]

mito-roGFP2-Orp1 in pLPCX (retroviral) [2]

cyto-Grx1-roGFP2 in pEIGW (lentiviral) [1]

Original probe references

[1] Gutscher et al. (2008) Real-time imaging of the intracellular glutathione redox potential. Nat Methods 5: 553-9

[2] Gutscher et al. (2009) Proximity-based protein thiol oxidation by H2O2-scavenging peroxidases. J Biol Chem 284: 31532-40

[3] Albrecht et al. (2011) In vivo mapping of hydrogen peroxide and oxidized glutathione reveals chemical and regional specificity of redox homeostasis. Cell Metab 14: 819-829

[4] Morgan et al. (2013) Multiple glutathione disulfide removal pathways mediate cytosolic redox homeostasis. Nat Chem Biol 9:119-125

[5] Albrecht et al. (2014) Redesign of genetically-encoded biosensors for monitoring mitochondrial redox status in a broad range of model eukaryotes. J Biomol Screen 19:379-86

[6] Morgan et al. (2016) Real-time monitoring of basal H2O2 levels with peroxiredoxin-based probes. Nature Chemical Biology 12: 437-443

Probe reviews

For detailed background information about roGFP2-based redox probes (mode of operation, limitations etc.):

Meyer and Dick (2010) Fluorescent Protein-based Redox Probes. Antioxid Redox Signal 13: 621-650

Ezeriņa et al. (2014) Imaging dynamic redox processes with genetically encoded probes. J Mol Cell Cardiol 73:43-49

Schwarzländer et al. (2015) Dissecting Redox Biology using Fluorescent Protein Sensors. Antioxid Redox Signal 24:680-712


For practical information on how to perform measurements (microscopy settings etc.) and data analysis:

Morgan et al. (2011) Measuring EGSH and H2O2 with roGFP2-based redox probes. Free Radic Biol Med 51: 1943-1951

Barata and Dick (2013) In Vivo Imaging of H2O2 Production in Drosophila. Meth Enzymol 526:61-82

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