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Fluorescein-Labeled DNA Probes for Homogeneous Hybridization Assays: Application to DNA E . coli Renaturation

Volume 51, Number 3 (March 1997) Page 401-406

Talavera, Eva M.; Alvarez-Pez, Jose M.; Ballesteros, Luis; Bermejo, Ruperto

The sensitivity of the emission intensity of fluorescein to environmental changes opens the possibility for developing homogeneous DNA assays. In this research, we were able to quickly and easily detect nucleic acid renaturation by using an E . coli DNA with approximately 4% conjugation of fluorescein to the N4-aminoethylmodified cytosine residues. We determined how the emission intensity of fluorescein attached to single-stranded DNA decreases when renatured to double-stranded DNA. The decrease in fluorescence efficiency is due to changes in the fluorescein monoanion-dianion ratio because of pH modifications in the surrounding of the fluorescein label. The decrease in fluorescence intensity was used for analysis of DNA E . coli renaturation kinetics.