Detection of DNA-protein binding in western blots by phosphorus-labeled and biotinylated DNA probes

Journal Article · Fri Apr 01 00:00:00 EST 1988 · Biotechniques; (USA)

OSTI ID:6749623

Herlt, M; Schwarz, H P; Neumann, R; Eggers, H J

Eukaryotic DNA-binding proteins can be detected by a filter binding assay combining protein blotting on nitrocellulose, incubation with DNA by filtration, and the application of radioactively or nonradioactively labeled DNA probes. Basic nuclear and non-nuclear standard proteins are assayed in dot blots as well as in Western blots from sodium dodecyl sulfate gels. The DNA-binding ability of fractionated proteins is compared employing two different blotting techniques, conventional electro-transfer and protein-renaturating capillary transfer. Biotinylated DNA probes exhibit high sensitivity and a distinct discrimination of detection signals corresponding only to defined DNA-binding proteins. In contrast, phosphorus-labeled DNA probes show higher sensitivity, but less effective resolving power, especially for bands localized close to each other. Using the DNA-incubation procedure described, biotinylated DNA probes are preferable to radioactively-labeled probes for screening DNA-binding proteins in complex protein fractions.

• OSTI ID: 6749623
• Journal Information: Biotechniques; (USA), Vol. 6:4; ISSN 0736-6205
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
PROTEINS
BIOASSAY
BIOCHEMICAL REACTION KINETICS
CATTLE
DNA
FILTRATION
PHOSPHORUS ISOTOPES
TRACER TECHNIQUES
ANIMALS
DOMESTIC ANIMALS
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
MAMMALS
NUCLEIC ACIDS
ORGANIC COMPOUNDS
REACTION KINETICS
RUMINANTS
SEPARATION PROCESSES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques