DNA concentration measurement

Chris LaRosa clarosa at biocomp.unl.edu
Sat Jun 17 11:46:52 EST 2000



elena wrote:
> 
> What if two stocks of DNA (concentration 15-30 µg/ml determined by spec)
> produce absolutely different standard curves when used to prepare standards
> within 0 - 0.001  µg/ml and read on microplate fluorometer?
> Results are consistent and repeatable. Linearity in any case is great.
> 
>
What kind of Fluorescent dye?? They are subject to different
interferences, and can bind differently to different types of dna.   
How different? 2 fold , 10 fold?
Either one is off or both are off... 
Also your fluorometer might be giving different gains at different
times. 



There are several possibilities....

You need to get a properly callibrated std DNA prep to start with. 
These can be bought.   You need to double check your homemade stocks. 
Perhaps run absorbance scans.

You may have interfering compounds in the buffer of one of the stocks.

You can always arbitrarily declare one of your stocks the "good" stock

The measurments may be off in absolute terms, but relative differences
may be good enough for your work.






More information about the Methods mailing list