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Storage of overexpressed protein


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#1 chn09

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Posted 10 November 2009 - 06:08 PM

Hi everybody,

Im working with protein purification and for that purpose ive stored my overexpressed protein (E. coli BL21, host) as cell pellets in -20 freezer for about 4 weeks. Is there any problem in storing overexpressed protein like this. or is storage of overexpressed protein in -70 most essential ?
pls help
thanks

#2 Vini

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Posted 10 November 2009 - 08:08 PM

Hi everybody,

Im working with protein purification and for that purpose ive stored my overexpressed protein (E. coli BL21, host) as cell pellets in -20 freezer for about 4 weeks. Is there any problem in storing overexpressed protein like this. or is storage of overexpressed protein in -70 most essential ?
pls help
thanks


Hi,
If i hv got u right, u hv stored the bacterial cell pellet in -20 and hv moved on to the lysis and furher purification steps?? In that case, its quite alright to store it at that temperature. Its only the protein which needs to be stored at -70 (and take care to add glycogen also)

#3 Prep!

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Posted 10 November 2009 - 08:35 PM

i second DRn.. IB's can be stored in -20 (but teh time has to be experimented) pure protein will have different storage temerature optimums.
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#4 chn09

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Posted 10 November 2009 - 09:49 PM

Hi
Thnks for ur reply. yes i have stored the overexpressed protein by centrifuging the cells in a falcon tube and stored it in -20 freezer. Now when i start my expt, i will have to thaw the cells in lysis buffer. at this time will the protein degrade or loose its activity?

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Posted 10 November 2009 - 10:00 PM

No it wont... but if its an IB, tehn u will have to refold it for obtaining the activity cause or else the prootein will be in the unfolded or misfolded form!!!
I understand you are gonna purify using downstream steps... in which sense u are doing just fine... hopefully you do have analytical techniques to qualify the identity of the protein!!
Support bacteria - They are the only culture some people have!!!
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