Measurement of in vivo DNA binding by sequence-specific transcription factors using UV cross-linking.
This paper describes an in vivo UV cross-linking protocol that is sensitive enough to detect DNA binding by sequence-specific transcription factors in Drosophila embryos and tissue culture cells. The strength of this approach is that it provides a quantitative measure of DNA binding in vivo with unambiguous identification of the factor involved in the binding. This assay often detects DNA binding properties of proteins that were not predicted from previous experiments, and it can be used to directly test diverse models of gene regulation in the context of a living organism.
Walter
J
J
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520-208114, USA.
Biggin
M D
MD
D000818:Animals; D001665:Binding Sites; D002458:Cell Fractionation / Q000379:methods; D002467:Cell Nucleus / Q000648:ultrastructure; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient / Q000379:methods; D003432:Cross-Linking Reagents; D004247:DNA / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D003902:Detergents; D004330:Drosophila; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004625:Embryo, Nonmammalian; D012521:Sarcosine / Q000031:analogs & derivatives; D013379:Substrate Specificity; D014157:Transcription Factors / Q000302:isolation & purification / Q000378:metabolism*; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D003902:Detergents; D014157:Transcription Factors; C025231:sarkosyl; D004247:DNA; D012521:Sarcosine