Calculating DNA Concentrations

[skillen]

Calculating DNA Concentrations

• Quote

Post by skillen » Sat Nov 28, 2009 2:07 am

Why must you calculate DNA concentrations by only using the dilutions that have a A260 nm value between 0.050 and 1.5? I've done the calculations, but I don't know WHY they need to be done that way. Any help is appreciated.

[JackBean]

• Quote

Post by JackBean » Sat Nov 28, 2009 12:55 pm

1) if you look at the whole spectra, you will see, that if is absorbance higher than 1, it is not straight line, but more like zigzag, so it's not much accurate
2) absorbance is log of the original light and the light gone through (A = I_0/I). So, imagine, if you have absorbance of 0.05 or 1.5, what part of light actually goes through? It is either more than 90% or less than 10% (in the case of 0.1 and 1, respectivelly, calculate real numbers for you case), so this is not much accurate either

[skillen]

• Quote

Post by skillen » Sat Nov 28, 2009 5:42 pm

Thanks very much for the help!