A Polarized Photobleaching Study of DNA Reorientation in Agarose Gels

Bethe A. Scalettar, Paul R. Selvin, Daniel Axelrod, Melvin P. Klein, John E. Hearst

Research output: Contribution to journalArticlepeer-review

Abstract

Polarized fluorescence recovery after photobleaching (pFRAP) has been used to study the internal dynamics of relatively long DNA molecules embedded in gels that range in concentration from 1% to 5% agarose. The data indicate that, even in very congested gels, rapid internal relaxation of DNA is largely unhindered; however, interactions with gel matrices apparently do perturb the larger amplitude, more slowly (microseconds to milliseconds) relaxing internal motions of large DNAs. The relationship between this work and recent studies which indicate that internal motions of DNA play an important role in the separation achieved with pulsed-field gel electrophoresis techniques is discussed. The polarized photobleaching technique is also analyzed in some detail. In particular, it is shown that “reversible” photobleaching phenomena are probably related to depletion of the ground state by intersystem crossing to the triplet state.

Original languageEnglish (US)
Pages (from-to)4790-4798
Number of pages9
JournalBiochemistry
Volume29
Issue number20
DOIs
StatePublished - May 1 1990
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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