1. What Is PCR Efficiency?

PCR efficiency measures how effectively DNA is amplified during each cycle of the polymerase chain reaction. An ideal efficiency of 100% means the DNA quantity doubles with each cycle.

2. How Does The Calculator Work?

The calculator uses the PCR efficiency equation:

Where:

• \( E \) — PCR efficiency (decimal)
• \( slope \) — Slope of the standard curve (Ct vs log(dilution))

Explanation: The equation converts the slope of your standard curve into a PCR efficiency percentage. Perfect efficiency (100%) corresponds to a slope of -3.32.

3. Importance Of PCR Efficiency

Details: Knowing your PCR efficiency is crucial for accurate quantification in qPCR experiments. Efficiency between 90-110% is generally acceptable.

4. Using The Calculator

Tips: Enter the slope value from your standard curve plot (Ct values vs log10 of template concentration). The slope is typically negative.

5. Frequently Asked Questions (FAQ)

Q1: What is an ideal PCR efficiency?
A: 100% efficiency means perfect doubling each cycle. 90-110% is generally acceptable for most applications.

Q2: What slope corresponds to 100% efficiency?
A: A slope of -3.32 corresponds to 100% efficiency.

Q3: Why might my efficiency be below 90%?
A: Possible causes include inhibitor presence, poor primer design, suboptimal reaction conditions, or template degradation.

Q4: Can efficiency be over 100%?
A: Yes, though uncommon. May indicate pipetting errors, contamination, or non-specific amplification.

Q5: How often should I check PCR efficiency?
A: With every new primer set and periodically thereafter, especially if changing reagents or instruments.