Jacob, You should be able to get rid of the problem within a few days as long as everyone in the lab is willing to cooperate.
Autoclaving works (it helps to use longer autoclave cycles - 40 minutes instead of the regular 20). If you confirm that phage is your issue - walk through the lab and carefully think about places where bacteria may be permanently colonized (the insides of shaker-incubators, sink traps, AC vents, sink borders, low-vacuum supply lines, condensate channels near autoclaves, etc.) and try to get these locations sterilized either chemically or physically. This would be a good time to dispose of old liquid media and suchlike. Plaque test representative samples of your competent cells (especially if you make your own) and glycerol stocks. Dry-heat the glassware and bleach the non-disposable plastic stuff. Inquire with the building manager regarding the pest population - are there loads of rats or mice traipsing through your lab at night? The key is to do this at the same time throughout the entire lab space. This can be tricky since most of us work in multi-user labs. Ultimately it's in everyone's best interest to get the lab phage-free so it should not be too difficult to convince people that a lab cleanout day is a good thing. Remind people that robust potty hygiene is more than a personal choice when they work in a biotechnology lab (it's remarkable that phage infection can both cause diarrhoea [by transmitting verocitoxin genes from bug to bug] and be the solution to diarrhoea [by lowering populations of enteropathogenic bacteria]). In the worst case scenario you may have to bribe/persuade people to get rid of potted plants and aquaria that may be present in the lab or the immediately adjoining offices - these moist environments may harbor phage-carrying bacteria. Good luck, Artem "Nothing is built on stone; all is built on sand, but we must build as if the sand were stone" Jorge Luis Borges -----Original Message----- From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of Jacob Keller Sent: Thursday, July 02, 2009 1:06 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Shredded E coli pellets Okay, it seems that the consensus is phage infection. Is there anything to seal the diagnosis? Also, does anybody have literature on de-phaging glassware? I am assuming that regular autoclaving will not do the trick? Jacob ******************************************* Jacob Pearson Keller Northwestern University Medical Scientist Training Program Dallos Laboratory F. Searle 1-240 2240 Campus Drive Evanston IL 60208 lab: 847.491.2438 cel: 773.608.9185 email: j-kell...@northwestern.edu ******************************************* ----- Original Message ----- From: "Jacob Keller" <j-kell...@md.northwestern.edu> To: <CCP4BB@JISCMAIL.AC.UK> Sent: Thursday, July 02, 2009 11:42 AM Subject: [ccp4bb] Shredded E coli pellets > Dear crystallographers, > > I recently expressed some new constructs, and found after my usual > expression protocol that the cell pellets were not compacted at the bottom > corner of the bottles us usual, but were instead smeared as a film on the > side, and further, were somewhat clumpy, like clots, and with a smaller > pellet in the usual location. The centrifugation was exactly as usual. I > noticed that there was also a bit more foam in the medium than usual, but > I am not convinced that this was the issue, although it might be a > symptom. My suspicion is that the constructs are lethal and cause cell > lysis, but I am not sure. Has anybody seen this phenomenon before, and > gotten to the bottom of it? > > Jacob Keller > > ******************************************* > Jacob Pearson Keller > Northwestern University > Medical Scientist Training Program > Dallos Laboratory > F. Searle 1-240 > 2240 Campus Drive > Evanston IL 60208 > lab: 847.491.2438 > cel: 773.608.9185 > email: j-kell...@northwestern.edu > ******************************************* >
