B-PER Lysis--CHEM 384
Ken Christensen
Abstract
The Thermo Scientific B-PER Bacterial Protein Extraction Reagent enables mild extraction of proteins from bacteria ( E. coli ) without the need for mechanical disruption. The reagent may be used for soluble protein extraction and inclusion body purification from bacterial cell lysates. The B-PER Reagent with Enzymes is supplied with lysozyme and DNase I to improve the extraction efficiency of large (> 70 kDa) molecular weight proteins and proteins expressed in inclusion bodies.
Steps
B-PER Lysis to extract soluble protein
Pellet bacterial cells by centrifugation at 5000 × g in a tared tube for 0h 10m 0s.
Decant your spent LB and determine the mass of your cell pellet.
Freeze your pellet in the -20°C freezer overnight or quickly freeze your pellet in liquid nitrogen. When you are ready to proceed, thaw your cells at room temperature.
Add 4mL of B-PER Reagent per gram of wet cell pellet. Pipette the suspension up and down until it is homogeneous.
Optional: Add 2µLof lysozyme and 2µL of DNase I per 1mL of B-PER Reagent added.
Incubate 0h 15m 0s at room temperature.
Centrifuge lysate at high speed (~15,000 × g) for 0h 5m 0s to separate soluble proteins from the insoluble proteins.
