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SDS-Page Gel Recipes

SDS-Page Gel Recipes[]

Brief Description

Seperating gel 5% 7% 9% 12.5% 15%

Protocol Details

Lower Gel

Reagents 5% 7% 9% 12.50% 15%
  2 minigels 2 minigels 2 minigels 2 minigels 2 minigels
30% Acrylamide mix 1.7 ml 2.4 ml 3.0 ml 4.2 ml 5 ml
4xLGB 2.5 ml 2.5 ml 2.5 ml 2.5 ml 2.5 ml
20% SDS 50 ul 50 ul 50 ul 50 ul 50 ul
Degas Water 5.7 ml 5.0 ml 4.4 ml 3.2 ml 3.2 ml
10% APS (0.1g/ml; fresh) 50 ul 50 ul 50 ul 50 ul 50 ul
TEMED (undil) 3.3 ul 3.3 ul 3.3 ul 3.3 ul 3.3 ul

 

Stacking Gel Buffer (SGB, 0.5 M Tris)
12.1 g Tris up to 200 ml of H2O, PH 6.8 (store in 4C)

Lower Gel Buffer (LGB, 1.5 M Tris)
36.3 g Tris, up to 200 ml of H2O, PH 8.8 (store in 4C)

20% Sodium Dodecyl Sulfate (SDS)
20 g SDS, up to 100 ml in H2O

1.5% Ammonium Persulfate (APS)
1.5 g APS, up to 100 ml in H2O (Aliquot and store in -20C)

 

Prepare Plates:

Ensure that everything is clean (Best way to ensure no leakage!)

Pour lower gel:

  1. Combine Acrylamide, water, SDS, LGB
  2. Mix by swirling (don't create bubbles)
  3. Add APS and TEMED (swirl to mix)
  4. Pipet mixture between plates to about 2/3 up with enough room for a few cm's from the bottom of the comb (move quickly)
  5. Add water or 70% EtOH to make top smooth.
  6. wait about 20 min.
  7. Pipet Upper Gel displacing water/EtOH to about 1-2cm from top.
  8. Insert comb displacing the rest of the water/EtOH.
  9. Let sit for about 15 min.

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