A nonradioactive method for detecting DNA-binding activity of nuclear transcription factors / 华中科技大学学报(医学)(英德文版)
Journal of Huazhong University of Science and Technology (Medical Sciences)
;
(6): 227-9, 2003.
Artigo
em Inglês
| WPRIM
| ID: wpr-635460
ABSTRACT
To determine the feasibility of a nonradioactive electrophoresis mobility shift assay for detecting nuclear transcription factor, double-stranded oligonucleotides encoding the consensus target sequence of NF-kappa B were labelled with DIG by terminal transferase. After nuclear protein stimulated with phorbol 12-myristate 13-acetate (PMA) or PMA and pyrrolidine dithiocarbamate (PDTC) electrophoresed on 8% nondenaturing poliacrylamide gel together with oligeonucleotide probe, they were electro-blotted nylon membrane positively charged. Anti-DIG-AP antibody catalyzed chemiluminescent substrate CSPD to image on X-film. The results showed that nuclear proteins binded specifically to the NF-kappa B consensus sequence in the EMSA by chemiluminescent technique method and the activity of NF-kappa B in PMA group was more than that in PMA + PDTC group. It is suggested that detection of NF-kappa B by EMSA with chemiluminescent technique is feasible and simple, which can be performed in ordinary laboratories.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Transcrição Gênica
/
Ativação Transcricional
/
Transativadores
/
NF-kappa B
/
Ratos Sprague-Dawley
/
Ensaio de Desvio de Mobilidade Eletroforética
/
Proteínas de Ligação a DNA
/
Medições Luminescentes
Idioma:
Inglês
Revista:
Journal of Huazhong University of Science and Technology (Medical Sciences)
Ano de publicação:
2003
Tipo de documento:
Artigo
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