How to express toxic proteins in bacteria - correction!

Emir Khatipov ekhatipo at NOSPAMmidway.uchicago.edu
Fri Sep 15 19:21:39 EST 2000


Thanks Peter, a really great post! Hope it will be as useful for everybody,
as it is for me.

I myself have a question, and would be happy to get a good advise. In my own
work I found that whereas BL21(DE3)pLysS cells are easy to lyse, it is
rather difficult to clear the lysate without spinning it down twice (once at
12000 x g (20') and then in the ultracentrifuge at ~40,000 x g), and still
the supernatant may contain greazy buoyant smear. The main inconvenience for
me is that when the ultracentrifugation is omitted, the lysate appears
cloudy, and seemingly contains chelators that cause sheering of cobalt from
Talon resin I used to purify His6-proteins. Is there any trick to eliminate
ultracentrifugation and the greazy stuff?
Emir

"Dr. Peter Gegenheimer" <PGegen at UKans.nolospamare.edu> wrote in message
news:7opiGDf98QgB-pn2-6qxUvBHWzdtY at rnaworld.bio.ukans.edu...
The previous post had some errors, so let me summarize the correct answer.
In
...







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