Preparing a SDS-PAGE Gel

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Separating Gel

For 12 mL final volume (2 gels)
Combine, adding APS and TEMED last
Pour into cassette, overlay with the top layer of water saturated butanol

Component	6%	7.5%	10%	12.5%
SDS-PAGE Separating Gel Buffer	3 mL	3 mL	3 mL	3 mL
Acrylamide Solution	2.4 mL	3 mL	4 mL	5 mL
Water	6.6 mL	6 mL	5 mL	4mL
APS (make fresh, 0.1g/mL in water)	200uL	200 uL	200 uL	200 uL
TEMED	30 uL	30 uL	30 uL	30uL

For a Phos-Tag Gel, add 24 uL 5 mM Phos-Tag and 24 uL 10 mM MnCl in the separating gel only

Stacking Gel

Rinse polymerized separating gel with water
For 10 mL (2 gels)
Combine, adding APS and TEMED last
Pour over separating gel and insert comb

Component
SDS-PAGE Stacking Gel Buffer	2.5 mL
Acrylamide Solution	1.7 mL
Water	4 mL
APS (make fresh, 0.1g/mL in water)	200 uL
TEMED	30 uL