RING1B-BMI1f was diluted to 1 μM final concentration in EMSA buffer (50 mM HEPES, pH 7.2, 100 mM NaCl, 10 mM MgCl2, 1 μM ZnCl2, 1 mM DTT, 5% glycerol, 0.01% BSA) and incubated with varying concentrations of compound at room temperature for 90 minutes. H2BOG488 nucleosomes were added to a 10 ng/μL final concentration in 10 μL reaction and reactions were incubated at room temperature for 30 minutes. 1 μL DNA loading dye was added to each reaction (Life Technologies). Reaction species were separated by electrophoresis at 4° C under native conditions using 7.5% Tris-glycine acrylamide gel (BioRad). Gels were imaged with Typhoon Trio+ 9410 Variable Mode Imager (GE Healthcare) and analyzed with ImageQuant TL 7.0 (GE Healthcare).
Typhoon trio 9410 variable mode imager
Manufactured by GE Healthcare
The Typhoon Trio+ 9410 Variable Mode Imager is a versatile laboratory instrument designed for the detection and analysis of a wide range of biomolecules. It utilizes multiple detection modes, including fluorescence, chemiluminescence, and phosphor imaging, to enable comprehensive analysis of various samples, such as gels, blots, and microplates.
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2 protocols using typhoon trio 9410 variable mode imager
1
RING1B-BMI1 Nucleosome Binding Assay
2
RING1B-BMI1 Nucleosome Binding Assay
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RING1B-BMI1f was diluted to 1 μM final concentration in EMSA buffer (50 mM HEPES, pH 7.2, 100 mM NaCl, 10 mM MgCl2, 1 μM ZnCl2, 1 mM DTT, 5% glycerol, 0.01% BSA) and incubated with varying concentrations of compound at room temperature for 90 minutes. H2BOG488 nucleosomes were added to a 10 ng/μL final concentration in 10 μL reaction and reactions were incubated at room temperature for 30 minutes. 1 μL DNA loading dye was added to each reaction (Life Technologies). Reaction species were separated by electrophoresis at 4° C under native conditions using 7.5% Tris-glycine acrylamide gel (BioRad). Gels were imaged with Typhoon Trio+ 9410 Variable Mode Imager (GE Healthcare) and analyzed with ImageQuant TL 7.0 (GE Healthcare).
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