Determination of the molecular weight of DNA-binding proteins using UV-crosslinking and SDS-PAGE.
We describe the use of UV-crosslinking in combination with SDS-PAGE to determine the approximate molecular weight of DNA-binding proteins. A 5-bromo-2'-deoxyuridine (5-BrdU)-substituted, radioactively labeled double-stranded oligonucleotide representing the protein binding site is incubated with a crude nuclear extract containing the protein of interest. Following irradiation with a UV light source, the DNA/protein complex is subjected to SDS-PAGE and its molecular weight determined by comparison with appropriate protein standards.
Wolf
S S
SS
Genetics and Microbiology Department, Norwich, UK.
Roder
K
K
Schweizer
M
M
D000818:Animals; D001973:Bromodeoxyuridine / Q000031:analogs & derivatives; D004256:DNA Polymerase I / Q000235:genetics; D004268:DNA-Binding Proteins / Q000032:analysis / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D008110:Liver Extracts / Q000032:analysis; D008970:Molecular Weight; D009687:Nuclear Proteins / Q000032:analysis; D051381:Rats; D012015:Reference Standards; D012967:Sodium Dodecyl Sulfate; D014466:Ultraviolet Rays
D004268:DNA-Binding Proteins; D008110:Liver Extracts; D009687:Nuclear Proteins; D012967:Sodium Dodecyl Sulfate; D004256:DNA Polymerase I; D001973:Bromodeoxyuridine