[Open_electroporator] culture shock docs -- culture_shock/LAB_MANUAL.md

[John Griessen]

I am confused by the following:

     repeat the following (3) times
         centrifuge the cell solution at the calculated RPM for 3 minutes
         discard the liquid, allowing the "cell pellet" to remain
         add 1mL distilled H2O
         resuspend by pulsing with vortexer

     resuspend once more in H2O, either 40µL or 200µL
         for 0.2 cm electroporation cuvette, pipet 40 µl
         for 0.1 cm electroporation cuvette, pipet 200 µl

     Transfer 1 pg–100 ng of plasmid DNA (1–5 µl) to cells and mix without vortexing.

Electroporation at low temperature (0-4 °C) seems to work best according to evidence, and efficiencies may drop ~100-fold at room 
temperature

     for best results, pre-chill the cuvette and cell suspension on ice for 1 to 2 minutes

     transfer cell solution to electroporation cuvette
         for 0.2 cm electroporation cuvette, maximum volume during pulse is 400 µl
         for 0.1 cm electroporation cuvette, maximum volume during pulse is 80 µl


Are the different volumes really recommended, or accidents?  I just looked closely at a 1mm cuvette
and see the top of the electrodes are insulated by plastic and there is a shorter well than the 2mm cuvette has.
The well between electrodes goes down 5mm at 10mm wide, then slopes in from the edges at 45 degrees
for 5mm.  So it has area 10x10 - (5x5) =75 sq mm and is 1mm across, so 75 microliters.  I'd say the
best way to fill these is aim for 75 ul with tolerance of + 10ul/- 0ul.  The overfill will not affect the
cuvette resistance, and may decrease yield by not getting fully zapped above the end of the electrode, but
the rest of what is filling it will get a consistent zap from the culture shock.