I eluted plasmid from zymopure miniprep kit which claims that endotoxin levels of ≤ 1EU/ug. This plasmid served as a control for my transfection for other test plasmids in murine embryonic stem cells. My test plasmids were eluted from the zymopure maxiprep kit which claims endotoxin levels of ≤ 0.025 EU/µg of plasmid DNA with optional EndoZero™ Spin-Column. Can I transfect my control plasmid got from miniprep and test plasmids obtained from the maxi kit for direct comparison of control and test plasmids irrespective of their source from mini and maxi kits? There is a difference of endotoxin levels between these kits.
It sounds like your endotoxin is low enough to not worry about it, unless you specifically study inflammatory signaling.
There are however other batch-to-batch plasmid characteristics that can have large effects on transfection efficiency. For a pivotal make-or-break question that goes into your publication, I think it would be best practices to prepare the experimental and control plasmids in identical ways. For routine research, in preliminary experiments (i.e. almost always), I think it can be OK to work with the different kits. You should always do a few routine tests, to make sure your plasmids are comparable, regardless of how they're prepared:
(1) Concentration should be at least somewhat close to each other.
(2) A260/A280 and A260/A230 ratios should be similar.
(3) Degree of supercoiling should be similar.
(4) Band intensity on gel electrophoresis should be similar, assuming identical A260 loading.
Thanks, John for the clarification. So batch-to-batch plasmid characteristics can have large effects on transfection efficiency!
Interesting question! As rightfully pointed out, the endotoxin levels in either of the preps is low enough to be ignored. Also, in my opinion if you're concerned about the difference, you could isolate plasmids using any one of the methods but same for both plasmid types. Yes batch-to-batch variation exists and could affect transfection result. One can include additional wash step (washing buffer) to achieve higher purity. Also, it shouldn't be a big issue to often isolate fresh plasmids for transfection and would even be considered as ideal by many!
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