Identification of Nucleic Acid Binding Proteins Using Nondenaturing Sodium Decyl Sulfate Polyacrylamide Gel Electrophoresis (SDecS-PAGE)

Abstract

Methods for the identification and characterization of nucleic acid binding proteins, such as DNA binding transcription factors, typically involve gel retardation assays (1,2) or Southwestern analysis (3). Gel retardation assays allow the detection of DNAbinding factors by assessing the degree to which protein binding affects the electrophoretic mobility of specific DNA sequences. One drawback of gel retardation assays is that only the presence of the protein is indicated; specific information concerning protein molecular weight, or other characteristics, is obtained only through additional methods. Southwestern analysis detects DNA binding proteins through the use of nucleic acid probes applied to protein blots prepared from sodium dodecyl sulfate (SDS) gels. The Southwestern technique relies on the limited ability of proteins to renature after SDS-gel electrophoresis and does not specifically identify binding by protein complexes, which are dissociated during sample processing.