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

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

Research output: Contribution to journalArticle

47 Citations (Scopus)

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

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Nucleic Acids
Gels
Experiments
Software
Motion Pictures
Practice Guidelines
Hydroxyl Radical
Nucleotides

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Semiautomated and rapid quantification of nucleic acid footprinting and structure mapping experiments. / Laederach, Alain; Das, Rhiju; Vicens, Quentin; Pearlman, Samuel M.; Brenowitz, Michael D.; Herschlag, Daniel; Altman, Russ B.

In: Nature Protocols, Vol. 3, No. 9, 2008, p. 1395-1401.

Research output: Contribution to journalArticle

Laederach, A, Das, R, Vicens, Q, Pearlman, SM, Brenowitz, MD, Herschlag, D & Altman, RB 2008, 'Semiautomated and rapid quantification of nucleic acid footprinting and structure mapping experiments', Nature Protocols, vol. 3, no. 9, pp. 1395-1401. https://doi.org/10.1038/nprot.2008.134
Laederach, Alain ; Das, Rhiju ; Vicens, Quentin ; Pearlman, Samuel M. ; Brenowitz, Michael D. ; Herschlag, Daniel ; Altman, Russ B. / Semiautomated and rapid quantification of nucleic acid footprinting and structure mapping experiments. In: Nature Protocols. 2008 ; Vol. 3, No. 9. pp. 1395-1401.
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