Koch Lab:Protocols/Microsphere-DNA tethering/Glass, dig, biotin, microsphere, 4kb DNA/BGB Prep
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Below are some old notes from Steve several years ago...they can probably be tweaked. For example, you can make more than 1.5 ml at a time.
To make 5 mg / ml BGB in popping buffer (BGB5pop) 1. Weigh out about 15 mg of BGB powder. 2. Add 1.5 ml of 2x popping buffer and mix by vortexing. 3. Run through 0.2μm syringe filter, using 3 ml syringe***. 4. Measure out volume that came through, and add equal amount water. Now should have 5 mg / ml BGB in 1x popping buffer (minus amount of protein that stuck to filter). (Can run Bradford assay to estimate concentration.)
Store at 4C (do not freeze), and can be used for a few weeks (keeping solution sterile). It’s not known how long it remains good. Also not known whether freezing affects blocking properties. If you determine is OK to freeze, then recipe can obviously be scaled up.
1x Popping buffer = 50 mM Sodium Phosphate pH 7.0, 50 mM NaCl, 10 mM EDTA, 0.02% Tween-20
- Note: Solubility of BGB (casein) varies widely with common buffers and differs for different commercial preparations. Solution should look clear, and should easily pass through 0.2 μm filter.