Sonication of cells in ultrasound bath, without probe - (Oct/16/2014 )
I am going to express and purify a recombinant protein in E. coli and only have an ultrasound bath to conduct the lyzis with (e.i. no sonicator with probe and no french press)
Is it possibe to lyze the cells efficiently in an ultrasound bath? (I have heard that it isn't). If so, could anyone give me some advice on how to do it and what to think of.
I don't want to do the sonication with only lyzosyme since it will become very expensive and I am of the oppinion that freeze thaw lyzis is not very reliable. Therefore I think that I have to go forward with the ultrasound bath.
Thankful for all help
I doubt that a bath sonicator will work, the strength of the ultrasound is too low. Could you try a detergent lysis?
I don't want to risk denaturing the protein.
So maybe I am left with freeze/thawing anyway. I am just affraid that it will not be efficient enough.
Not to say something you don't want to hear, but if you want to get into the business of protein expression and purification, you really should find a sonicator or french press, these are pretty much the only reliable ways to lyse (bacterial) cells and keep proteins natured. Trying to do it without either is like trying to do molecular biology without a PCR machine.
Though there are some proprietary chemical options like Bug Buster (Millipore), but never tried them myself.
You "could" express your protein with a periplasmic signal sequence so that it gets sent to and folds in the periplasm, in which case you can get at your protein by osmotic shock. But yields are very poor, and if your protein of interest isn't naturally a peripasmic protein it is unlikely to fold proper. This is usually only done when all else fails. I'd rather fork over the $$'s for a sonicator than deal with that headache though...