| Solution Information | help | |
| Enzyme: | DNA dC->dU-editing enzyme APOBEC-3A | |
| inhibitor: | BDBM62590 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Assay Materials: Protein dilution buffer: Tris (pH 7.4) at 15 mM, NaCl at 150 mM, Triton X-100 at 0.5%, glycerol at 10% Oligo dilution buffer: TE at 20 mM Oligo: 5' d FAM-AAATATCCCAAAGAGAGA-TAMRA 3': BioSearch Technologies UDG: New England Biolabs A3A-MycHis: University of Minnesota 1N NaOH 2M Tris (pH 7.9) Assay plate: Corning 1536 Well Black Solid Bottom Plate (Catalogue # 3724) I. Compound Addition: 1. Dose response curves contained 10 concentrations of compounds obtained using a twofold serial dilution. Compounds were serially diluted in 100 % DMSO (1% final DMSO concentration). Using LabCyte Echo, transfer 40 nL compounds in triplicate into assay plate columns 5 - 48. Transfer 40 nL of DMSO to positive and negative control wells in columns 1 - 4. 2. Centrifuge plates at 1000 rpm for 1 min. 3. Seal the plates and leave them at RT. II. Set up of APOBEC3A assay: 4- Prepare protein dilution buffer 5- Prepare oligo dilution buffer 6- Prepare intermediate A3A at 0.4 ng/ul. T | |
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