Hello everybody, I’m posting this arguably off topic somewhat related to previous query question on behalf of a colleague:
Does anyone have experience with Origami cells growing in fermenter? Many searches yield nothing as if these cells have never been used to grow in place other than flasks. Vaheh Oganesyan www.medimmune.com From: CCP4 bulletin board [mailto:[email protected]] On Behalf Of Artem Evdokimov Sent: Thursday, March 19, 2015 6:25 PM To: [email protected] Subject: Re: [ccp4bb] Protein expression Question: you're taking a 3-ml equivalent out of 500 ml culture, and processing it as if it was a 3ml culture? Or are you basing the result on processing the entire 500ml? Reason I ask this is simply to make sure your extraction/purification timing is the same in both cases. It matters! Assuming that the protein really is not soluble at 500ml and is somewhat soluble at 3ml - which expression set up are you using? IPTG or autoinduction? more details is better. For example, if you use IPTG - how do you determine when to induce - OD? or some other measure? The advice to change aeration is solid. Try it both ways, meaning up or down - it could go either way depending on the difference in your set up. A 3ml culture in a 24-well block at 800 rpm is MUCH better aerated than 500ml culture in non-baffled 1L flask at 250rpm. You also may want to play with temperature, and if you're not using AIM - try it out. Artem - Cosmic Cats approve of this message On Thu, Mar 19, 2015 at 2:33 PM, Reza Khayat <[email protected]<mailto:[email protected]>> wrote: Hi, We can express quite a bit of soluble protein when growing 3ml cultures. However, the protein becomes insoluble (inclusion bodies) when we scale up to 500ml cultures. Has anyone experienced such a problem, and found a solution to it? Thanks. Best wishes, Reza Reza Khayat, PhD Assistant Professor Department of Chemistry City College of New York New York, NY 10031 http://www.khayatlab.org/ 212-650-6070<tel:212-650-6070> To the extent this electronic communication or any of its attachments contain information that is not in the public domain, such information is considered by MedImmune to be confidential and proprietary. This communication is expected to be read and/or used only by the individual(s) for whom it is intended. If you have received this electronic communication in error, please reply to the sender advising of the error in transmission and delete the original message and any accompanying documents from your system immediately, without copying, reviewing or otherwise using them for any purpose. Thank you for your cooperation.
