IUBio

DNA-Standards

Thomas Pfitzner Thomas.Pfitzner at medizin.uni-koeln.de
Wed Mar 17 08:28:20 EST 1999


Hi,
I have a few questions regarding DNA-Standards for quantitative PCR. We
have cloned our target sequence into a plasmid. What would you suggest:
Using native or linearised Plasmid for standards ?
How do you produce your linearised Standard ?
(We tried gel elution using QuiaExII, which seem'd to work)
Do you recommend Carrier DNA and which type ? (We used Salmon Sperm DNA
from Stratagene which had a profound negative effect on PCR-reaction)
Which method of DNA-Quantification do you prefer ? (What about your
experience with OD at 260 nm compared to other, more sophisticated methods?)
How about long-term storage of your standards ? Does this work, or do you
prefere to produce a new set of standards after a period of time ?
Thank you very much.



Thomas Pfitzner



Dr. Thomas Pfitzner
Laboratory of Immunotherapy
University of Cologne
LFI, E4, R703
Josef Stelzmann Stra=DFe 9
50924 Cologne
Germany
Tel.: +49-(0)221-478-4418 / -5993
=46ax.: +49-(0)221-478-6383
email: Thomas.Pfitzner at medizin.uni-koeln.de

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