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E. coli cell lysis for Protein Isolation URGENT HELP PLEASE

Dr Mick Jones m.d.jones at ic.ac.uk
Fri Jan 14 11:23:08 EST 2000


Hi

I am running a student practical next week, and I no longer have access
to a sonicator for making cell lysates from E. coli.

The experiment simply involves growing E. coli containing a pGEX fusion
construct, lysing the cells, and analysing the cleared lysate on an
SDS/PAGE gel and staining with Coomassie blue.  We also use the lysate
to run on another SDS/PAGE and western blot and probe with a monoclonal
antibody against the GST protein.

The experiments work very well.

This time round I do not have access toa sonicator to lyse the E. coli
cells.

I usually work with a 2 ml culture of E. coli.

CAN ANYBODY HELP?

What I need is a simple method for lysing the cells that doesn't use a
sonicator.

It would be nice if it is simply a chemical disruption with relatively
simple lab raegents, as opposed to a commercial reagent.

The proteins we are looking at are soluble.

Any help and advice, protocols greatfully accepted

Many thanks

Please reply directly to my email address:  m.d.jones at ic.ac.uk

Mick Jones
Lecturer in Molecular Virology
Department of Infectious Diseases and Microbiology
Imperial College School of Medicine
Hammersmith Hospital, Du Cane Road, London, W12 0NN
Tel:  020-8383-3328;  Fax:  020-8383-3394
(Note new London code)
email:  m.d.jones at ic.ac.uk
-------------------------------------------
"To Infinity and Beyond"  Buzz Lightyear (Toy Story)
"Crackin' Toast, Gromit"  Wallace (The Wrong Trousers)

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