User:Tamra L. Fisher/Notebook/Experimental Biological Chem/2012/04/09

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Objective

Make a protein gel.

Description

1st Attempt at Making Gel Today

  1. Make the resolving gel:
    • 1.65mL H2O
    • 2.0mL 30% acrylamide mix
    • 1.25mL 1.5M Tris (pH 8.8)
    • 0.05mL 10% SDS
    • 0.05mL 10% ammonium persulfate (add this right before pouring the gel)
    • 0.002mL TEMED (add this right before pouring the gel)
  2. Pour the gel between the short plate and spacer plate (after they have been secured in the casting frame on the casting stand). Squirt a layer of methanol on top of the resolving gel for a straight, level layer of gel.
  3. This layer of gel never polymerized, so I need to try again.

2nd Attempt at Making Gel Today

  1. Make fresh 10% APS (0.1g ammonium persulfate + 1mL distilled H2O)
  2. Make new 200mL 1.5M Tris, pH 8.8 (36.34g Tris base + H2O + HCl (to pH))
  3. Make the resolving gel:
    • 1.65mL H2O
    • 2.0mL 30% acrylamide mix
    • 1.25mL 1.5M Tris (pH 8.8)
    • 0.05mL 10% SDS
    • 0.05mL 10% ammonium persulfate (add this right before pouring the gel)
    • 0.002mL TEMED (add this right before pouring the gel)
  4. Pour the gel between the short plate and spacer plate (after they have been secured in the casting frame on the casting stand). Squirt a layer of methanol on top of the resolving gel for a straight, level layer of gel.


Notes

Neither resolving gel solidified today. I will have to continue to troubleshoot this problem tomorrow.



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