1. What is QPCR Efficiency?

QPCR efficiency (E) is a measure of how efficiently the target DNA is amplified during each cycle of the quantitative PCR reaction. It is calculated from the slope of the standard curve.

2. How Does the Calculator Work?

The calculator uses the QPCR efficiency equation:

Where:

• \( E \) — QPCR efficiency (dimensionless)
• \( slope \) — Slope of the standard curve

Explanation: The efficiency is derived from the slope of the standard curve plot (Ct vs. log input DNA). An ideal reaction has a slope of -3.32, corresponding to 100% efficiency.

3. Importance of QPCR Efficiency

Details: Efficiency values between 90-110% are generally acceptable. Values outside this range may indicate problems with primer design, PCR conditions, or standard curve quality.

4. Using the Calculator

Tips: Enter the slope value from your standard curve. The slope should be negative (typically between -3.1 to -3.6 for acceptable efficiency).

5. Frequently Asked Questions (FAQ)

Q1: What is an ideal QPCR efficiency?
A: 100% efficiency is ideal, corresponding to perfect doubling of product each cycle. 90-110% is generally acceptable.

Q2: What causes low QPCR efficiency?
A: Common causes include poor primer design, PCR inhibitors, suboptimal reaction conditions, or degraded template.

Q3: What causes efficiency >100%?
A: This may indicate pipetting errors, contamination, or non-specific amplification.

Q4: How often should efficiency be checked?
A: Efficiency should be determined for each primer pair and checked periodically (with each new batch of reagents).

Q5: Can I use this with SYBR Green or probe-based assays?
A: Yes, the efficiency calculation works for both detection methods.