How do I determine the efficiency of my Real-Time PCR assay?

/How do I determine the efficiency of my Real-Time PCR assay?

How do I determine the efficiency of my Real-Time PCR assay?

2018-01-24T08:51:04+00:00December 4th, 2017|

The simplest and most commonly used method is the dilution or standard curve method. This method calculates PCR efficiency using the linear regression slope of a dilution series based on the following equation:

E = 10(-1/slope) -1

The ideal slope is -3.32, which correlates to an amplification efficiency of 100%, meaning exactly one copy per cycle. Slopes in the range of -3.60 to -3.10 are generally considered acceptable for Real-Time PCR. These slope values correlate to amplification efficiencies between 90% (1.9) and 110% (2.1). The correlation coefficient or R2 value should be at least 0.985 or higher.

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