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How can I have same plateau fluorescence?


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#1 egerecske

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Posted 07 July 2009 - 11:59 PM

Dear all, I want to do a melting curve analysis with equal amount of different type of DNA of a mixed reaction (I mean I will mix the two DNA before the reaction). To get nice results, IŽd need to know what starting amount is necessary to reach the plateau with the same fluorescence level. I tried to dilute samples, but I still have differences, and I found they not necessarily correlate with sample Ct values. What factors influence the fluorescence of the plateau? How can I change it?
any suggestions are appreciated.
e.

#2 molgen

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Posted 08 July 2009 - 06:36 AM

When I do real-time I have good duplicates but rarely do I get the same plateau.

pH can effect the fluorescence as well as yield.

What I can't understand is why do you want the plateau? What info can or do you expect to get from it?




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