samples (from yeast)= Whole cells, spheroplasts, crude mitochondria, and isolated mitochondria
I am not getting your question .... but with PCR products we cannot go for this because the DNA gets diffused...
Hi, DNA/RNA would diffuse quite fast in an agarose gel (a few hours). I don't know about putting whole cells etc but I have a feeling it would not run well, even if they didn't diffuse. If you are talking about polyacrylamide gels, they would diffuse as well, but I'm not sure how fast.
Hi there,
The purpose of electrophoresis is to generate sharp bands. If you leave thee sample in the well overnight it will diffuse naturally into the gel. And I don't see the point loading the samples without running the electrophoresis right after. Either you run the gel overnight with reduced voltage or you load the samples in the morning and run the normal voltage.
Thanks all. It is a polyacrylamide gel; I should've been more clear that this is the electrophoresis step of a western blot.
In fact, you have a porous body(your gel), so DNA or protein can move through it slowly.
Electric current just made it faster and straight.
So it does not matter if its whole cell extracted or small DNA, with time it will start diffuse out, max to max 30 min. If you run on next day you will not have anything in the gel its the same goes for SDS-PAGE as well. So prefer run as soon as possible after loading sample into gel. One thing you do, you can ran sample at low voltage, (But overnight running is too much).
Nope. It will diffuse out of the porus materix of the gel with that much time. The glycerol in the loading buffer will allow it to settle to the wells bottem, but with this much time it will go to equilibrium and difuse taking your samplw with it.
NO! You can pour your gel the night before, write down exactly how you want to load your sample and even mix in the loading dye in advance. But you will lose your samples if you load them in advance. This is not a shortcut you can use.
Szanowny Panie Profesorze,
Dlaczego Pańskie pytania i dyskusja znalazły się w dziale, w którym mowa o kryształach, nanokryształach i przemianach fazowych w kryształach? CZy nie powinny byc w innym miejscu w RG?
Pozdrawiam,
Paweł Tomaszewski
You could reduce the voltage to make the gel run slower, especially with the 20x20 cm2 gels. Then you can fix them first thing in the morning. In that case, I'd use a gradient gel to limit diffusional band-spreading (assuming you are talking about protein electrophoresis).
No dear! I wonder why would u do such a thing.
U can very well make your gel in the evening and keep it in fridge for overnight (even for two consecutive nights), and then load your sample followed by applying some volatge to it.
If u will load your sample and then let it sit for overnight, high chances are it will diffuse with buffer.
In case u r performing such experiment (as you queried), and u succeed, let us know!
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