PCR problems using Streptomyces cosmid DNA as template?

M.D. Farrar MICMDF at leeds.ac.uk
Tue Feb 29 09:06:36 EST 2000


In article <89e09j$5q8$1 at willow.cc.kcl.ac.uk>,
   Colin Dolphin <colin.dolphin at kcl.ac.uk> wrote:
>Dear All
>
>
>I'm attempting to amplify a predicted gene in Streptomyces coelicolor
>using the appropriate cosmid (approx. 30k) DNA, containing the gene
>sequence, as template without, so far, any success. I was wondering
>whether this could be due to the high GC content of the template. Would
>fragmenting the cosmid with an appropriate RE help and/or the use of an
>additive such as DMSO? 
>
>
>Colin Dolphin
>Dept of Pharmacy
>Kings College London
>London SE1 8WA
>colin.dolphin at kcl.ac.uk


Hi Colin.

I had problems with PCR using Streptomyces coelicolor genomic DNA until I used 
10% (v/v) DMSO in the reaction - it worked a treat!

Good luck


Mark Farrar
Division of Microbiology
School of Biochemistry and Molecular Biology
University of Leeds
Leeds, LS2 9JT
micmdf at leeds.ac.uk




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