Quantifying double-strand breaks and clustered damages in DNA by single-molecule laser fluorescence sizing

Elena M. Filippova, Denise C. Monteleone, John G. Trunk, Betsy M. Sutherland, Stephen R. Quake, John C. Sutherland

Research output: Contribution to journalArticle

24 Scopus citations

Abstract

Fluorescence from a single DNA molecule passing through a laser beam is proportional to the size (contour length) of the molecule, and molecules of different sizes can be counted with equal efficiencies. Single-molecule fluorescence can thus determine the average length of the molecules in a sample and hence the frequency of double-strand breaks induced by various treatments. Ionizing radiation-induced frank double-strand breaks can thus be quantified by single-molecule sizing. Moreover, multiple classes of clustered damages involving damaged bases and abasic sites, alone or in combination with frank single-strand breaks, can be quantified by converting them to double-strand breaks by chemical or enzymatic treatments. For a given size range of DNA molecules, single-molecule sizing is as or more sensitive than gel electrophoresis, and requires several orders-of-magnitude less DNA to determine damage levels.

Original languageEnglish (US)
Pages (from-to)1281-1290
Number of pages10
JournalBiophysical Journal
Volume84
Issue number2 I
DOIs
StatePublished - Feb 1 2003

ASJC Scopus subject areas

  • Biophysics

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