Supplementary Figure 2: Fluorescent switching through cleavage of disulfide conjugate dye on readout probes is highly efficient. | Nature Methods

Supplementary Figure 2: Fluorescent switching through cleavage of disulfide conjugate dye on readout probes is highly efficient.

From: Profiling the transcriptome with RNA SPOTs

Supplementary Figure 2

(a) 20 rounds of hybridization are accomplished by extinguishing fluorescent signals through reduction of disulfide conjugated dye to readout probes using TCEP, followed by re-hybridization of next unique secondary readout probes. (b) Both priming regions (grey in the probe schematic) used in synthesizing gene specific primary probes are also used as a registration marker through the hybridization of Alexa 488 conjugated readout probes. Majority of the fluorescent spots stay even after 20 rounds of hybridizations. The amide bond between the Alexa 488 dye (shown in yellow) and primer readout probes used as a registration marker is not affected by TCEP. (Scale bars: 2μm.) (c) The fluorescent signals in each channel after treatment of 50mM of TCEP for 5 minutes at room temperature is reduced to minimal to none. (Scale bars: 5μm.)

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