title: Examination of DNA-binding activity of neuronal transcription factors by electrophoretical mobility shift assay.
abstract: Electrophoretical mobility shift assay (EMSA) is a simple, rapid, and highly sensitive technique for detection of single- or double-stranded DNA-binding proteins such as transcription factors in crude nuclear extracts (F.M. Ausubel, R. Brent, R.E. Kingston, D. D. Moore, J.G. Seidman, J.A. Smith, K. Struhl (Eds.), Current Protocols in Molecular Biology, Greene Publishing Associates and Wiley-Interscience, 1989, pp. 12.0.1-12.2.10 [1]; J. Carey, Gel Retardation. Methods Enzymol., 208 (1991) 103-117 [2]). By using this technique, it is possible to quantify the abundance, relative affinity and binding specificity of DNA-binding proteins. Since proteins which bind specifically to radiolabeled DNA probes retard the mobility of the probe during electrophoresis (it also called gel retardation assay), discrete bands correspond to the individual DNA-protein complexes. Furthermore, EMSA allows one to determine which member(s) of a certain protein family are included in the DNA-protein complex by means of specific antibodies raised against the DNA-binding protein (supershift assay).
authors.affiliation: Ishida Group of Clock Gene, National Institute of Bioscience and Human Technology, Agency of Industrial Science and Technology, MITI, 1-1, Higashi, Tsukuba-Science City Ibaraki 305, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Kako
authors.forename: H
authors.initials: H
authors.lastname: Wakamatsu
authors.forename: T
authors.initials: T
authors.lastname: Hamada
authors.forename: M
authors.initials: M
authors.lastname: Banasik
authors.forename: K
authors.initials: K
authors.lastname: Ohata
authors.forename: T
authors.initials: T
authors.lastname: Niki-Kuroiwa
authors.forename: S
authors.initials: S
authors.lastname: Suzuki
authors.forename: J
authors.initials: J
authors.lastname: Takeuchi
authors.forename: N
authors.initials: N
authors.lastname: Ishida
mesh_terms: D000818:Animals; D001667:Binding, Competitive; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D008297:Male; D009474:Neurons / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein