Fluorescein-12-dUTP could be used in directly enzymatic labeling of DNA/cDNA, such as PCR, random primed DNA labeling reaction, primer extension, Nick Translation and others. It can be incorporated as a substitute for its natural counterpart dTTP. The Dye-labeled DNA/cDNA probes acquired are ideally suited for fluorescence hybridization applications - FISH or microarray-based gene expression profiling. Optimal substrate properties and labeling efficiency is ensured by an optimized linker attached to the C5 position of uridine. Recommendatory fluorescein-12-dUTP/dTTP ratio for PCR and Nick Translation: 30-50% Fluorescein-12-dUTP/ 70-50% dTTP. Please note: Please protect the Dye-labeled dUTP from exposure to light and experiment under low light conditions. The optimal final concentration of the Dye-labeled dUTP might very depending on the practical application and assay conditions. For generating optimal product yields and high incorporation rates, an individual optimization for the Dye-labeled-dUTP/dTTP ratio is recommended.
Fluorescein-12-dUTP is used as a substrate for:
- Taq polymerase
- phi29 DNA Polymerase
- DNA Polymerase I (holoenzyme and Klenow fragment)
- Terminal Transferase
- Reverse transcriptase (e.g., from AMV and M-MuLV).
References:
[1]. Wiegant et al. (2001) Probe Labeling and Fluorescence in situ Hybridization. Current Protocols in Cytometry 8:8.3.1.
[2]. Yie SM1, Xiao R, et, al. Progesterone regulates HLA-G gene expression through a novel progesterone response element. Hum Reprod. 2006 Oct;21(10):2538-44. Epub 2006 May 9.