1. What is PCR Efficiency?

PCR efficiency (E) is a measure of how effectively DNA is amplified during each cycle of the PCR reaction. An ideal PCR has 100% efficiency, meaning the amount of product doubles with each cycle.

2. How Does the Calculator Work?

The calculator uses the PCR efficiency equation:

Where:

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

Explanation: The slope of the standard curve is used to calculate the efficiency of the PCR reaction. The ideal slope for 100% efficiency is -3.32.

3. Importance of PCR Efficiency

Details: PCR efficiency affects the accuracy of quantitative PCR results. Efficiency between 90-110% is generally considered acceptable for reliable quantification.

4. Using the Calculator

Tips: Enter the slope value from your standard curve plot. The value should be negative (typically between -3.0 and -3.6 for good PCR efficiency).

5. Frequently Asked Questions (FAQ)

Q1: What is the ideal PCR efficiency?
A: The ideal efficiency is 100%, which corresponds to a slope of -3.32. This means the DNA doubles perfectly with each PCR cycle.

Q2: What slope value indicates good PCR efficiency?
A: A slope between -3.1 and -3.6 typically indicates good efficiency (90-110%).

Q3: What causes poor PCR efficiency?
A: Common causes include poor primer design, suboptimal reaction conditions, PCR inhibitors, or template degradation.

Q4: How can I improve PCR efficiency?
A: Optimize primer design, adjust Mg2+ concentration, use high-quality reagents, and ensure proper thermal cycling conditions.

Q5: Should I use this for absolute quantification?
A: For absolute quantification, you need a standard curve with known concentrations and efficiency between 90-110%.