We have three shaking incubators (15°C below ambient to 80°C) for protein expression, which can each hold up to 6 x 1 litre cultures. We are soon to receive an Eppendorf BioFLo120 for larger scale protein production runs.

Expression

We currently use the Shuffle T7 express cells (NEB) strain of E. coli for protein expression. These cells are an excellent expression strain, offering increased solubility of proteins with di-sulphide bonds. The range of E. coli expression cells offered will be expanded based on collective user needs, if you require a different strain, please get in touch.

Following each transformation of a plasmid into the E. coli expression cells we carry out a test for expression and solubility. This is undertaken by 18’C incubation overnight in 0.5 mM IPTG, after which the cells are lysed and expression and solubility are analysed by SDS-PAGE.

If soluble protein is detected expression conditions can either be further optimised or protein expression can be scaled up.

We currently have 3 incubators, with a capacity to hold 5 x 1 litre cultures per incubator. We can provide either sterile LB or TB media for this purpose. Expression can either be overseen by the user or be run directly by the Biotechnology Facility technical staff at no extra cost.

A nanodrop 2000C is available for monitoring cell growth if required.

Cell lysis

The method of cell lysis used is dependent on samples size.

For analysis of very small sample volumes, e.g. for test expression samples for SDS_PAGE, we recommend using the Bugbuster.

For medium scale cell lysis, e.g. for use prior to purification, we recommend the sonicator.

For larger samples volumes we have a cell disruptor available.