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q PCR efficiency

q PCR efficiency - Real-Time PCR and Quantitative PCR Forum

q PCR efficiency - Real-Time PCR and Quantitative PCR Forum


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  #1  
Old 08-22-2008, 10:05 AM
Pipette Filler
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Default q PCR efficiency



Hi Everyone. First time for me to write in such forum. I am now doing qPCR (learning actually) and I have a question to anyone who might be able to help.

I am using the standart curve method to determine my primers efficiency. It works like this:
- I am making series of standart dilutions (i.e. 1:5)
- than the efficiency is calculated in % from the slope of the curve that I get (i.e. 95%).

The thing is that I am not sure above which value of PCR efficiency is my experiment considered 'good'. Is it ok if the efficiency is about 90 % ? Or does it have to be 100 % ? Can I use the primers under the same conditions if the efficiency calculated by this method happens to be below 90 % ?

thanks !

Ivan
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Old 08-25-2008, 04:43 PM
Pipette Filler
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Default Re: q PCR efficiency

Hi,
how are you calculating efficiency? what values are you plugging in the equation to get your percentages?
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Old 08-27-2008, 02:53 PM
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Default Re: q PCR efficiency

hi, thanks for the will to help. actually, the software calculates the efficiency from the slope of the standart curve (made as plot of Ct (cycle thereshold) versus log of starting quantity of template).

E% = 10(-1/slope)-1*100

Last edited by Ivan; 08-27-2008 at 03:54 PM.
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Old 09-05-2008, 01:38 PM
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Default Re: q PCR efficiency

Hi Ivan,
usually I consider as good efficency a value between 85 and 100%. The slope between 3.3 and 3.5...... I hope it will be useful.
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