Re: [DIYbio] Re: Designing a DIY Gene Electroporator

On Fri, Oct 18, 2013 at 10:17 AM, John Griessen <john@industromatic.com> wrote:
> On 10/18/2013 04:01 AM, Nathan McCorkle wrote:
>>
>> What about the issue of finding uF sized caps for 1.8-2.5kV? Is there
>> a way to simulate a cap's discharge? Use inductors or something?
>
>
> If you mean stretch out a pulse, yes, inductance will do that, and/or
> oscillate, so you have to get specific.

No I meant to substitute for these seemingly hard-to-find capacitors.
Eliminating them from the design.

>
> And yes, computer simulations are good for fine tuning power designs --
> if you know what you want as a goal. MLP said she was rebooting work
> on an electroporator/melaminometer the other day...
>
> Probably, with observations from your piezo
> spark generator experiments, and some more simple tests, we could skip
> reverse engineering brands like Gene Pulser and such and avoid patent
> attacks.
> There's probably no patent on basic function though, just features like
> arc suppression. The high resistive DI water method seems easiest and
> lowest cost parts, since it requires the least power flow. I find a newish
> product uses an optional module the size of a book adding big capacitors for
> long "square wave" pulses to solutions with < 1k Ohm
> cuvette resistance. Square wave is a separate system to make -- put off for
> now.
>
> If common practice in convenient/costly lab gear is to provide an
> exponential decay
> from capacitor discharge of 5, 10, 25 uF, and yet, multiple pulses are just
> fine and OK,
> why not keep costs low by using a 2 or 5 uF cap only and repeating while
> necessary?

I read many short pulses don't get the job done, or the cells have a
much higher death rate or something.

>
> One reason might be that HV makes radio interference, and it's best to make
> one big zap
> then be quiet and say, "Who, me?" The design of this kind of thing needs
> a shielding metal box to keep interference low, and when sold as a product,
> it needs FCC
> compliance testing as an interference source. When an arc gets started, it
> can
> generate heat and you can get a steam explosion, or just a blast of gases
> when in air,
> so that's another reason for a solid container around the cuvette.
>
> Leaving some of the hard parts for later, wouldn't a small capacitor fired
> as many times as it takes
> be as good as a bigger one for the energy for a HV pulse? Does polarity
> matter in electroporation?

I believe the commercial E.coli unit I've used did single pulse per
cuvette/reaction tube. Polarity would matter in the sense that it's
going to be driving the DNA via electrophoresis... the field polarity
probably doesn't matter as much for pore formation. I bet the
difference between single long pulses and many short is that long DNA
will take some time to electrphorese from solution into the cytoplasm
through an open pore, so pulsing it will essentially reset the whole
system each time, not giving the DNA enough time to slip into and all
the way through the open membrane.

> Obviously, it can't matter from the start, since orientation of bugs in a
> solution is random, but
> after a discharge drops, what if it reverses some, giving your bugs a few
> wiggles of AC before dying out?

Probably doesn't matter for single pulses.

> That will come naturally for a capacitor discharging into and inductive
> transformer to up the voltage
> to 2kV.
>

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