Welcome to the new version of CaltechAUTHORS. Login is currently restricted to library staff. If you notice any issues, please email coda@library.caltech.edu
Published January 9, 1996 | Published
Journal Article Open

DNA-protein binding assays from a single sea urchin egg: A high-sensitivity capillary electrophoresis method

Abstract

A capillary electrophoresis method has been developed to study DNA-protein complexes by mobility-shift assay. This method is at least 100 times more sensitive than conventional gel mobility-shift procedures. Key features of the technique include the use of a neutral coated capillary, a small amount of linear polymer in the separation medium, and use of covalently dye-labeled DNA probes that can be detected with a commercially available laser-induced fluorescence monitor. The capillary method provides quantitative data in runs requiring < 20 min, from which dissociation constants are readily determined. As a test case we studied interactions of a developmentally important sea urchin embryo transcription factor, SpP3A2. As little as 2-10 x 10^6 molecules of specific SpP3A2-oligonucleotide complex were reproducibly detected, using recombinant SpP3A2, crude nuclear extract, egg lysates, and even a single sea urchin egg lysed within the capillary column.

Additional Information

© 1996 National Academy of Sciences. Contributed by Eric H. Davidson, September 28, 1995. We thank Chiou-Hwa Yuh, James A. Coffman, Helen Lee, and Chieh Chang for their efforts in preparation and provision of the samples and reagents that we used. This work was supported by the Beckman Institute of California Institute of Technology, by National Institutes of Health Grant HD-05753, and by a subcontract from National Science Foundation Science and Technology Grant BIR9214821. J.X. was supported by National Institutes of Health Training Grant HD-07257. The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Attached Files

Published - PNAS-1996-Xian-86-90.pdf

Files

PNAS-1996-Xian-86-90.pdf
Files (960.9 kB)
Name Size Download all
md5:2695399ff0b6b2e42547a3fabfbf070b
960.9 kB Preview Download

Additional details

Created:
August 22, 2023
Modified:
October 18, 2023