In article <53tt16$9gc at info.service.rug.nl>
g.j.e.j.hooiveld at farm.rug.nl (Guido Hooiveld) writes:
> Hi everybody!
>> After performing a prep I estimate the amount and purity of my plasmid DNA sample by measuring
> the A260 and A280. I use the ratio of A at 260 and 280 nm to get an indication of the purity of
> my sample.
>> I was told an A280/A260 ratio of 1.8 to 1.9 indicates a highly purified DNA preparation and its
> theoretically limit is 2.0. However, I often find a ratio larger than 2.0; like 2.1 to 2.2. So,
> is this (theoretically) value of 2.0 correct, if yes, does anybody have a suggestion why I find
> values >2; if not, what is this value?
>> And what about the factor you have to use to determine the concentration of DNA present:
> multiply A260 with 50 to obtain the concentration (ug/ml), or use 35? Which value is correct?
>> Any comments would be appreciated!
>> Thanks in advance,
>> Guido
>> --
> ***************************************************************************
> * Guido Hooiveld | *
> * Groningen University Center for Pharmacy |phone: (+)31 50 3637565 *
> * dep. of Pharmacokinetics and Drug Delivery | (+)31 50 3633272 *
> * A. Deusinglaan 1 |fax: (+)31 50 3633247 *
> * 9713 AV Groningen | *
> * The Netherlands | *
> ***************************************************************************
>
The value of 50 ug/ml per OD is for double strand DNA and 35 is for
single strand DNA.
The proper ratio is 260/280, not 280/260. Recalculate and see what you
have.
Michael Benedik
Department of Biochemical Sciences
University of Houston
benedik at uh.edu
