ESA009 at ED.SAC.AC.UK ("Dr Rob Harling") wrote:
>We're using RAPD-PCR to study diversity in epiphytic Pseudomonas
>species. We prepare the template DNA using Flowgen's Puregene kit,
>which is basically an SDS cell lysis, ammonium acetate protein
>precipitation, RNAse, isopropanol DNA precipitation, then dissolve in
>water. The resulting DNA is of good spectrophotometric quality.
>PCR works if we use freshly - prepared DNA, ie extracted from the
>bacteria then run on the same day. If we store the DNA at 4C for
>only one week, then PCR fails, we get zilch in the lanes at any
>concentration of template. The DNA still looks the same after one week
>when run on a gel, ie there doesn't seem to be nuclease breakdown
>of the DNA. Does anyone know what's happening to the DNA in storage?
What is your storage buffer (if any) and why do you store at 4C?
>Thanks
>Rob (still got my hair) Harling
>Dr Rob Harling
>SAC (Scottish Agricultural College)/
> University of Edinburgh
>West Mains Road
>Edinburgh EH9 3JG
>Scotland, UK
>tel: +44 (0)131 535 4000
>fax: +44 (0)131 667 2601
>e mail: esa009 at ed.sac.ac.uk
