The low-concentration, 5 pg/ul standard will take many more cycles to cross the same threshold – and therefore the Ct will be higher. The high-concentration, 50 ng/ul standard will cross the detection threshold first, generating a “low” Ct. The Ct values are inversely proportional to the concentration of DNA in the standards.The Ct value is what ultimately is used to create the standard curve. When the fluorescent signal crosses the detection threshold the cycle number is recorded as a Ct value, or threshold cycle value. During each PCR cycle, the amount of fluorescent signal for each standard in the dilution seies is measured.In this example, the standards consist of a 10-fold dilution series ranging from 50 ng/ul down to 5 pg/ul. Everyone wants to know how much DNA is in their extract, but then they ask: how can I tell if my estimate is accurate?Ī standard curve is a tool that allows us to estimate the DNA concentration of unknown samples by comparing them to standards with known DNA concentrations.
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