Nucleic Acids Research, Vol 24, Issue 22 4535-4542, Copyright © 1996 by Oxford University Press
D Proudnikov and A Mirzabekov
Several procedures have been described for fluorescent labeling of DNA and
RNA. They are based on the introduction of aldehyde groups by partial
depurination of DNA or oxidation of the 3'-terminal ribonucleoside in RNA
by sodium periodate. Fluorescent labels with an attached hydrazine group
are efficiently coupled with the aldehyde groups and the hydrazone bonds
are stabilized by reduction with sodium cyanoborohydride. Alternatively,
DNA can be quantitatively split at the depurinated sites with
ethylenediamine. The aldimine bond between the aldehyde group in
depurinated DNA or oxidized RNA and ethylenediamine is stabilized by
reduction with sodium cyanoborohydride and the primary amine group
introduced at these sites is used for attachment of isothiocyanate or
succinimide derivatives of fluorescent dyes. The fluorescent DNA labeling
can be carried out either in solution or on a reverse phase column. These
procedures provide simple, inexpensive methods of multiple DNA labeling and
of introducing one fluorescent dye molecule per RNA, as well as
quantitative DNA fragmentation and incorporation of one label per fragment.
These methods of fluorophore attachment were shown to be efficient for use
in the hybridization of labeled RNA, DNA and DNA fragments with
oligonucleotide microchips.
ARTICLES
Chemical methods of DNA and RNA fluorescent labeling
Engelhardt Institute of Molecular Biology, Moscow, Russia.
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