measure the DNA concentration

[Silvia_84]

Hi all,

I should measure the DNA concentration I obtained after the DNA purification. I have a total volume of DNA solution of 100 ul and I would like to use the spectrophotometer. Since I have the 10mm cuvettes, how can I handle the volume of DNA to test in the cuvette? I mean, the reading measures will be done on 1 ml of solution into the cuvette, won't they? Should I dilute my DNA sample in water in order to fit the solution in the cuvette and measure the concentration ? If yes, how do I diluite it?

Thank you in advance.

[CharonY]

Depending on the lightpath you may be able to use ~500-700 ul. But either way, your DNA extract has to be diluted first. Typically the same buffer to resuspend the DNA is used, though practically water is alright in almost all cases. With regard to "how", what problems do you have?

[Silvia_84]

Yes, sorry. I was just asking what type of dilution is better in terms of quantity.. if to do 1:10 in water or other..sorry I am new with this stuff.

[CharonY]

That is your judgement call. It depends on how much you have extracted, how much you need for your analyses (as you do not want to waste all your sample) and the volume you need. So if you use 1:10 you will need 50-100 ul of your sample.

Also, there are 100 ul cuvettes that one can use to minimize required volume.

[Arete]

I have both a Qubit fluorometer and a Nanodrop spectrophotometer in my lab for quantification. Which device do you have? The nanodrop takes a 1ul sample, so I generally just quantify a stock extraction. The Qubit seems a little more consistent, but is more involved to use.