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electrophoretic mobility-shift assay (EMSA) was performed as described previously ( Tsui et al . 2006 ). The double-stranded DNA fragment containing the putative ChoRE (5′-TGTGCACGAGGGCAGCACGAGCCTC-3′), was 5′-end labeled with γ-P 32 ATP using T4 polynucleotide

′-GTTCCCTGTCGCTCTGTTtttGCTTATCTCTCCCGCCTT-3′ M10 −103/−100 bp S: 5′-CTGTCGCTCTGTTCCCGCccgTCTCTCCCGCCTTTTCGG-3′ M11 −98/−96 bp S: 5′-TCGCTCTGTTCCCGCTTATCctcCCCGCCTTTTCGGCGCCACC-3′ EMSA WT −121/−83 bp S: 5′-CTGTCGCTCTGTTCCCGCTTATCTCTCCCGCCTTTTCGG-3′ MT −102/−98 bp S: 5

). Electrophoretic mobility shift assay (EMSA) Preparation of nuclear extracts, probe labelling, super shifts and the general procedures for the EMSA analysis were conducted as described ( Mao et al. 2002 ). To generate the double-stranded probe, we used

, suggesting a cross-talk between the GLIS3 and GLI signaling pathways ( Kim et al . 2003 ). To identify the specific binding site for GLIS3, Beak and coworkers combined PCR and electrophoretic mobility shift assay (EMSA) to screen a mixture of 60 bp

Electrophoretic mobility shift assays (EMSA) were carried out with wild-type and mutant PITX2 and the 32 P-labeled double-stranded oligonucleotide 5′-ACCAGGATGC TAAGCC TGTGTC-3′, containing the CE3 PITX-specific binding site (in bold) of the proopiomelanocortin

and 7), while the mutated oligonucleotide had limited effect (lanes 4 and 8). Figure 6 Both endogenous and in vitro translated GATA proteins bind to position −101 of the rat LHβ gene promoter. (A) Electrophoretic mobility shift assay (EMSA

E-box at positions −10 to −5 relative to the transcriptional start. Bearing in mind that Usf2 transactivates numerous genes by binding to E-boxes ( Corre & Galibert 2005 ), electrophoretic mobility shift assays (EMSA) were performed with nuclear

). Electrophoretic mobility shift assay Recombinant GATA4 wild-type and mutated proteins were in vitro translated using a QuickCoupled TNT in vitro transcription and translation system (Promega). Electrophoretic mobility shift assays (EMSA; DNA-binding) were

's improved minimal essential medium was obtained from Cellgro-Mediatech, Inc. (Manassas, VA, USA). Fetal bovine serum (FBS) was from Equitech-Bio, Inc. (Kerrville, TX, USA). Oligonucleotide primers and EMSA nucleotide duplex probes were obtained from

EMSA and effects of α-helical (9–41) CRH, SB203580 (10 μM) and PD98059 (20 μM). (B) Supershift assay using antibody against the p50 subunit of NF-κB confirmed that the Ucn-stimulated increase in binding resulted from NF-κB activity in the nuclear