Gene Copy Number Calculator

Calculate DNA Copy Number for qPCR Experiments

The Gene Copy Number Calculator is an essential tool for molecular biologists performing quantitative PCR (qPCR) and other nucleic acid quantification assays. It allows you to convert the mass of a DNA sample (in nanograms) into the absolute number of DNA molecules (copies). Precise knowledge of copy number is critical for creating standard curves and determining the absolute quantification of target genes.

How to Calculate Gene Copy Number

The calculation is based on the relationship between the mass of the DNA, the length of the template, and the average weight of the nucleotides. The formula used to determine the number of copies is derived from Avogadro's constant and the molar mass of the DNA.

The formula is:

Where:

• Amount (ng): The total mass of the DNA sample.
• Length (bp): The size of the DNA template in base pairs.
• K (Weight Factor): The average molecular weight of a base pair. For double-stranded DNA (dsDNA), this is approximately 660 g/mol. For single-stranded DNA (ssDNA), it is 330 g/mol.
• 6.022 × 10²³: Avogadro's number (molecules/mole).
• 10⁹: Conversion factor from nanograms to grams.

Why Use This Calculator?

In standard curve generation for absolute quantification, you must dilute your template DNA to specific copy numbers (e.g., 10⁷, 10⁶, etc.). Spectrophotometers provide readings in ng/µL, which measures mass, not molecule count. Since a short DNA fragment weighs significantly less than a plasmid or genomic DNA, 100 ng of a PCR product contains far more copies than 100 ng of a whole plasmid. This calculator normalizes these differences to give you exact molecular counts.

Factors Affecting Accuracy

Ensure your inputs are precise:

• Template Purity: Contaminants (RNA, proteins) can inflate the perceived DNA mass, leading to overestimation of copy numbers.
• Length Accuracy: Ensure the base pair count includes the full plasmid or amplicon length being measured, not just the target gene sequence.
• Strandedness: Select the correct type (dsDNA vs ssDNA) as single-stranded cDNA or viral DNA has half the molecular weight of double-stranded DNA per unit of length.