We mainly use endotoxin-free kit for plasmid preparation due to disturbance of LDS.
But there are also non-endotoxin-free kit.
Why do we use it?
I think that non-endotoxin-free kit is cheaper than endotoxin-free kit if we use gram-positive bacteria as host of plasmid.
Are there other answers about this? Please answer in detail!!
Hello Kim!
In my experience the different usage depends on the purpose of the plasmid:
I would use the Non-Endotoxin-Free Kit for plasmids that you use in a non cell culture background (e.g. PCR template, Bacterial transformation, Cloning).
For mammalian cell lines or other susceptible cell lines I would use a Endotoxin-Free Kit to avoid contamination or negative effect on your precious cells. For example when I was working with insect cell lines I always used a Endotoxin-Free Kit to avoid contamination of my culture due to toxins or other impurities.
So basically the plasmid preparation is cleaner, therefore minimizing the risk of side effects while transforming the plasmid into your host cells.
Best regards,
Pascal
Hello Kim!
In my experience the different usage depends on the purpose of the plasmid:
I would use the Non-Endotoxin-Free Kit for plasmids that you use in a non cell culture background (e.g. PCR template, Bacterial transformation, Cloning).
For mammalian cell lines or other susceptible cell lines I would use a Endotoxin-Free Kit to avoid contamination or negative effect on your precious cells. For example when I was working with insect cell lines I always used a Endotoxin-Free Kit to avoid contamination of my culture due to toxins or other impurities.
So basically the plasmid preparation is cleaner, therefore minimizing the risk of side effects while transforming the plasmid into your host cells.
Best regards,
Pascal
Dear Kim
I will add to Pascal's excellent answer by saying that muco polysaccharides and proteoglycans are difficult to purify away with standard non endotoxin kits (essentially silica fines) resulting in lower purity DNA. Furthermore, if these complex molecules bleed through into your final DNA prep apart from deleterious effects on cells (in the case of endotoxin) they can also inhibit downstream enzymes in vitro
Thus like Pascal for various reasons outlines, purity, enzyme inhibition endo toxic shock to cells I would use an endotoxin purification kit with Gram positive bacteria
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