XFD546, manufactured by AAT Bioquest, is a bright orange fluorescent dye that is structurally similar to Alexa Fluor™ 546 (Thermo Fisher), offering high photostability and intense fluorescence. When conjugated to uridine triphosphate (UTP), XFD546-UTP functions as an alternative to natural UTP in T7 RNA polymerase-mediated in vitro transcription, enabling the synthesis of fluorescently labeled RNA probes. These probes are highly suited for multicolor fluorescence-based applications, including dual-color expression profiling, microarrays, fluorescence in situ hybridization (FISH), chromosome identification, whole chromosome painting, karyotyping, and gene mapping.

XFD546-UTP efficiently incorporates into RNA transcripts through T7 RNA polymerase, imparting robust fluorescence for the visualization and quantification of RNA localization, distribution, and dynamics in live or fixed-cell systems. With excitation and emission maxima at 561 nm and 572 nm, respectively, XFD546-UTP-labeled RNA is readily detectable under fluorescence microscopy and gel electrophoresis without additional post-staining. Furthermore, XFD546-UTP facilitates single-molecule fluorescence resonance energy transfer (smFRET) experiments, allowing real-time monitoring of co-transcriptional RNA folding and structural dynamics at the molecular level. The exceptional photostability and narrow emission profile of XFD546-labeled nucleotides make them highly advantageous for multiplexed fluorescence imaging, enabling precise signal discrimination in complex biological samples.