PCR template concentration - (May/19/2010 )

Hi there,

I am running a QPCR and with sybr green and I'm not sure how much template to use. I am going to titrate my template but am not sure what to use for my high-end titration point. My template stock concentration is 100um. Any advice?

Thanks!

Use 10e8 or 10e7 copies as highest concentration. Then dilute in 10-fold steps.

I'm sorry but I don't know what you mean....

Moksha on May 19 2010, 05:08 PM said:

I understood the exponential terms, what I don't understand is what is meant by "copies".

One copy is one molecule of template DNA.
What kind of template you have? Plasmid? cDNA? PCR product?
What is your template concentration? I don't think it is 100 uM. You were talking about primers probably.

Hi, if you are not sure how much of the template youwant to use, why not perform a 10-fold serial dilution of your template to check not only the efficiency of your run but also the good concentrations you can work with in succeeding runs. By the way, what knd of template are you working with? same questions as the one posted by vladoo.