User:Elaine Marie Robbins/Notebook/CHEM-496/2011/10/25

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Objective

To run an SDS-PAGE to determine whether or not MBP-intein fusion proteins are being expressed cells comparable to the cells frozen on 09/28/11.

Description

  1. Make solutions for a 12% discontinuous gel using the BioRad Mini-PROTEAN instructions.
  2. Set resolving gel solution in gel holder with methanol on top to make the top of the gel flat.
  3. Allow gel to sit for 20 min.
  4. Remove methanol.
  5. Add stacking gel solution.
  6. Load gels into SDS-PAGE.
  7. Load ladder and protein solutions into gel.
  8. Run SDS-PAGE.
  9. Stain gels with Coomassie Brilliant Blue R250 solution.
    1. Dissolve 0.2555 g Coomassie Brilliant Blue R250 in 90 mL of methanol:water (1:1 v/v) and 10 mL of glacial acetic acid.
    2. Filter solution through a Whatman No. 1 filter.
  10. Place on shaker overnight.

Data

Protein visibly traveled on gel after SDS-PAGE.

Notes

Protein solutions used were from group 2: