A Peculiar Event In Tranformation

Chris Boyd chrisb at hgu.mrc.ac.uk
Fri Jun 20 09:43:51 EST 1997


Paul Dunman (dunman at njmsa.UMDNJ.EDU) wrote:
: Randy Willis <willis at gandalf.psf.sickkids.on.ca> writes:

: >Paul N Hengen wrote:
: >> But, will they be able to grow under non-selective conditions? I think the
: >> cells undergo more stress (not less) at 4C. I used to keep all my slants of
: >> E. coli at 4C until I found out they last much longer if stored at room temp.
: >> Then again, who needs the cells when you're studying plasmids?  I now just
: >> make sure to keep any plasmid DNA I'm interested in at -20C.


: >Paul,

: >You may be correct.  My use of the word "stress" is very hand-waving.  I
: >do agree that it is best to keep a solution of DNA at -20C, and I do.  I
: >wonder what peoples' experiences have been with using cells taken from
: >stocks which have been frozen at -70C.  We maintain stocks, but
: >typically this is in case we run out of plasmid DNA and NOT for the
: >purposes of growths/expressions.  Any suggestions or ideas?

: >Best of luck to all in their endeavours...Randy


: Our lab typically stores all strains at -20C & -70C in 40% gylcerol.  
: I've had trouble recessitating (sp) strains from -20C after they've been 
: stored for several years.  However, I've never had trouble recessitating 
: strains from -70C.  I directly plate on selective media & "knock on wood" 
: haven't ran into a problem yet.  I should also mention, all plasmids are 
: also purified & stored @ -20C...just in case.

It is safer to store strains at -70C in at least 15% glycerol: some
strains last indefinitely at -20C, but others do not --  typically
those strains you try to retrieve six months later.

On the other hand, plasmid DNA, if sufficiently pure and concentrated
(and not too large), will last indefinitely at 4C.  I recently cut out
and subcloned a fragment from a plasmid stock I made in 1984 with no
problem whatsoever.

Best wishes,
--
Chris Boyd                       | from, | MRC Human Genetics Unit
chrisb at hgu.mrc.ac.uk             |  not  |  Western General Hospital
http://www.hgu.mrc.ac.uk/~chrisb |   for |   Edinburgh EH4 2XU, SCOTLAND



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