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

被引：58

作者：

Laederach, Alain ^{[1
,3
]}

Das, Rhiju ^{[4
]}

Vicens, Quentin ^{[5
]}

Pearlman, Samuel M. ^{[6
]}

Brenowitz, Michael ^{[7
]}

Herschlag, Daniel ^{[8
]}

Altman, Russ B. ^{[2
,3
]}

机构：

[1] Wadsworth Ctr, Dept Dev Genet & Bioinformat, Albany, NY 12208 USA

[2] Stanford Univ, Dept Bioengn, Stanford, CA 94305 USA

[3] Stanford Univ, Dept Genet, Stanford, CA 94305 USA

[4] Univ Washington, Dept Biochem, Seattle, WA 98195 USA

[5] Univ Colorado, Dept Chem & Biochem, Boulder, CO 80309 USA

[6] Stanford Univ, Biomed Informat Program, Stanford, CA 94304 USA

[7] Albert Einstein Coll Med, Dept Biochem, Bronx, NY 10461 USA

[8] Stanford Univ, Dept Biochem, Stanford, CA 94305 USA

来源：

NATURE PROTOCOLS | 2008年 / 3卷 / 09期

基金：

美国国家卫生研究院;

关键词：

D O I：

10.1038/nprot.2008.134

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

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 similar to 20 min. Although SAFA was developed to analyze hydroxyl radical (center dot 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.

引用

页码：1395 / 1401

页数：7

共 27 条

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