Basic Subcloning Question

Dr. Duncan Clark Duncan at nospam.demon.co.uk
Mon Jun 7 06:45:38 EST 1999


In article <37595FC6.62CE942F at ibm.net>, Nicolas Lemee <Lemee at ibm.net>
writes
> Again at 135$ a bag of
>Biorad cuvettes (not to mention the electroporator itself), I'd rather do heat
>shock and buy top quality cells.

Of course you could save all the money and make your own electroporator.
Straightforward to do as per a very old Biotechniques article (don't ask
me the year or author!) using fairly low voltage and very small gaps.
Mine uses 340V (240V AC rectified via an isolation transformer). Charge
a 22uf capacitor and discharge that across a gap created using two
appropriately insulated large aluminium blocks with a gap created by a
few layers of high voltage insulating tape (i.e. equivalent of
12500V/cm). By initially adjusting the gap and the capacitance one can
get very close to commercial machines for E.coli. You need cells in 20%
glycerol rather than 10%. Zap, overlay with 1ml SOC and remove cells. To
sterilize wipe the blocks with 70% EtOH and use again and again. Polish
as is required every 20 or so zaps (high voltage side only) with
automotive chrome cleaner. 

Duncan   
-- 
The problem with being on the cutting edge is that you occasionally get 
sliced from time to time....

Duncan Clark
DNAmp Ltd.
Tel: +44(0)1252376288
FAX: +44(0)8701640382
http://www.dnamp.com
http://www.genesys.demon.co.uk



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