Semiautomated and rapid quantification of nucleic acid footprinting and structure mapping experiments

Alain Laederach, Rhiju Das, Quentin Vicens, Samuel M. Pearlman, Michael Brenowitz, Daniel Herschlag, Russ B. Altman

Research output: Contribution to journalArticle

51 Scopus citations

Abstract

We have developed protocols for rapidly quantifying the band intensities from nucleic acid chemical mapping gels at single-nucleotide resolution. These protocols are implemented in the software SAFA (semi-automated footprinting analysis) that can be downloaded without charge from http://safa.stanford.edu. The protocols implemented in SAFA have five steps: (i) lane identification, (ii) gel rectification, (iii) band assignment, (iv) model fitting and (v) band-intensity normalization. SAFA enables the rapid quantitation of gel images containing thousands of discrete bands, thereby eliminating a bottleneck to the analysis of chemical mapping experiments. An experienced user of the software can quantify a gel image in ∼20 min. Although SAFA was developed to analyze hydroxyl radical (·OH) footprints, it effectively quantifies the gel images obtained with other types of chemical mapping probes. We also present a series of tutorial movies that illustrate the best practices and different steps in the SAFA analysis as a supplement to this protocol.

Original languageEnglish (US)
Pages (from-to)1395-1401
Number of pages7
JournalNature Protocols
Volume3
Issue number9
DOIs
StatePublished - 2008

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

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