plasmid standard curve qPCR

Severin, Ina via methods%40net.bio.net (by I.Severin from nioo.knaw.nl)
Wed Jan 7 05:06:43 EST 2009


Hi all,

I am currently working on the quantification of gene copy numbers and
transcripts and got to the point where I need to make my standard curve.
I have already isolated and linearized the plasmids containing the
insert of interest but am (already for a while) wondering for what exact
reason the linearization needs to be done. Unfortunately, I did not find
much information in the literature (at least not in ecological
research). Do the different formations of a plasmid behave differently
in a qPCR reaction? And in how far could the buffer used for the digest
interfere with my qPCR reaction? Do I need to clean my plasmid solution
(and if yes, what's the best way)?

 

Many thanks in advance!

Ina

 

 



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