Distilled water for Electrocompetent Cell Preparation
Posted 27 April 2011 - 08:50 AM
I have also read that some people use 1 mM HEPES buffer pH 7 to prepare electrocompetent cells as well. However, I have also heard that it makes no difference to the final transformation efficiency whether one uses distilled water or HEPES.
Is the distilled water okay because everything is kept so cold or something?
Posted 28 April 2011 - 08:39 PM
Posted 01 May 2011 - 10:16 PM
It works just fine, don't worry
Edited by leelee, 01 May 2011 - 10:16 PM.
Posted 02 May 2011 - 03:56 AM
Posted 05 May 2011 - 01:54 PM
It works fine for E. coli, but not for other species that are osmotically sensitive. These species often use an electroporation buffer of 1 mM HEPES, pH 7.5 plus 280 mM sucrose, chilled.
Hmmm. . .do you think there would be any benefit to providing the bacteria with a nice pH as well. Perhaps the electroporation efficiency would be a little higher? I guess this is nothing to fret about since just about everyone uses water with e. coli anyway.
I should also add that using HEPES instead of water seems to decrease the time constant when I electroporate, which may be a bad thing. Perhaps I need to change the voltage I electroporate with when I use HEPES. Since I don't know enough about this, I may be better off using the standard distilled water method. I would like to get the highest electroporation efficiency possible though since I am making a cDNA library.
Edited by azim58, 05 May 2011 - 02:01 PM.
Posted 05 May 2011 - 05:14 PM