Gene expression is regulated in part by protein transcription factors that bind target regulatory DNA sequences. Predicting DNA binding sites and affinities from transcription factor sequence or structure is difficult; therefore, experimental data are required to link transcription factors to target sequences. We present a microfluidics-based approach for de novo discovery and quantitative biophysical characterization of DNA target sequences. We validated our technique by measuring sequence preferences for 28 Saccharomyces cerevisiae transcription factors with a variety of DNA-binding domains, including several that have proven difficult to study by other techniques. For each transcription factor, we measured relative binding affinities to oligonucleotides covering all possible 8-bp DNA sequences to create a comprehensive map of sequence preferences; for four transcription factors, we also determined absolute affinities. We expect that these data and future use of this technique will provide information essential for understanding transcription factor specificity, improving identification of regulatory sites and reconstructing regulatory interactions.
|Number of pages||6|
|State||Published - Sep 2010|
Bibliographical noteFunding Information:
P.M.F. was supported by a Howard Hughes Medical Institute/Helen Hay Whitney Foundation Postdoctoral Fellowship. J.L.D., S.R.Q. and this work were supported by the Howard Hughes Medical Institute. We thank A. Potanina for assistance with fabrication of microfluidic devices, O. Homann for implementation of PSAM functionality with MochiView and D. Breslow, F. Caro, S. Churchman, M. Dimon, T. Kiers, A. Kistler, C. Nelson, K. Sorber, E. Yeh and I. Zuleta for careful reading of the manuscript.