|- The liquid medium is aspirated from the Petri dishes. A proteinase inhibitor cocktail (Complete; Roche) is added to the Petri dishes and the dishes are frozen at −80°C.|
|- Qproteome Bacterial Protein Prep kit (Qiagen) is added to each Petri dish. The whole content is transferred into a glass test tube and vortexed (10×30s cycles with 1 min breaks on ice). The beads and cell debris are removed by centrifugation (5 min at 5000g at 4°C) and are rewashed with lysis buffer.
- The proteins precipitation and resuspension protocol is the one provided by the publication.
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2 years ago
Author: Hollie Fowler
I am using 8M Urea to lyse my cells but after centrifugation my pellet is very viscous. This causes me trouble when I try to collect the supernatant. Is there any way to avoid it and do you think it will have an effect on the concentration of my protein?
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