bch0047 at uabdpo.dpo.uab.edu
Tue Feb 14 15:10:55 EST 1995
sjwhite at ksu.ksu.edu (Susan Jane White) wrote:
> I'm purifying genomic DNA from a filamentous fungus. I want to know how much
> I have, but I suspect I have too little (or of too bad quality) to get good
> spec readings. I tried using EB and spotting onto Saran wrap on a lightbox
> (ref. Sambrook et al.); this seemed to work OK, but the second time my standard
> curve was pretty much all the same - I'm not sure if they got concentrated by
> evaporation in the fridge or what. Does anyone have any easy, reliable method?
> I've seen ads for a "Dipstick" DNA assay (Invitrogen?) but it was pretty
> expensive and they wouldn't send me a sample (hmpf...). Are they nice? Is there
> an easy "home-made" version?
We have had problems with our spectrophotometer, so we had
to find a way to estimate our DNA concentrations without it for
a while. What we did was take a sample of isolated DNA and run
it on a gel to get an estimate of the amount and purity of the
isolated DNA. I have also tried using the DNA Dipstick kit.
It works fairly well if you only process 2 to 4 samples at a
time. It uses a cuvette as the reaction vessel, so I do 2 at
a time and face the sticks back to back. The sensitivity is claimed
to be as low as 0.1 ng/microliter. The difficulty I had with this
kit was actually esitimating to my satisfaction the amount of DNA
I had. The color makes it difficult to read in between 0.1 ng/microliter
and 0.5 ng/microliter. The ranges are: 0.1, 0.5, 1, 5, and greater
than 10 ng/microliter. Also, this kit does not give you an idea
of the purity of your DNA. The cheapest thing to do is to run a minigel
with small wells (for us, that's what we do).
I hope this helps.
The Gene Therapy Program
University of Alabama at Birmingham
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