2 replies to this topic

[miBunny]

In the DNA isolation protocol for using TriZol or TriReagent, the pellet is washed with trisodium citrate/10% ethanol. Does anyone know the purpose of the trisodium citrate in this step?

This has been bugging me... the only thing I can think of is that the DNA is pretty cruddy and a good amount of washing is needed. The trisodium citrate keeps the pH somewhat alkaline and 10% ethanol is enough to keep the DNA from solubilizing during the long wash step.

Does anyone have the real answer?

[DrRob]

miBunny, on Nov 7 2009, 02:20 AM, said:

In the DNA isolation protocol for using TriZol or TriReagent, the pellet is washed with trisodium citrate/10% ethanol. Does anyone know the purpose of the trisodium citrate in this step?

This has been bugging me... the only thing I can think of is that the DNA is pretty cruddy and a good amount of washing is needed. The trisodium citrate keeps the pH somewhat alkaline and 10% ethanol is enough to keep the DNA from solubilizing during the long wash step.

Does anyone have the real answer?

Sodium acetate is added to DNA prior to ethanol precipitation where the sodium ions serve to neutralize the negative (PO4) charges of the DNA backbone, essentially making the DNA less hydrophilic and more likely to precipitate. I suppose the tri-Na serves the same purpose here?

[miBunny]

Ohhhh I like that logic!

Each molecule will donate 3 sodiums so you will not need to add to much to the solution to neutralize the backbone which means the pH won't get too high.

Thanks!