Measuring sequencing accuracy

Understand quality scores, or Q scores, and what they mean for your sequencing run

Sequencing Quality Scores

Sequencing quality scores measure the probability that a base is called incorrectly. With sequencing by synthesis (SBS) technology, each base in a read is assigned a quality score by a phred-like algorithm1,2, similar to that originally developed for Sanger sequencing experiments.

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Sequencing Technology Video

See how Illumina SBS works.

The sequencing quality score of a given base, Q, is defined by the following equation:

Q = -10log10(e)

where e is the estimated probability of the base call being wrong.

  • Higher Q scores indicate a smaller probability of error.
  • Lower Q scores can result in a significant portion of the reads being unusable. They may also lead to increased false-positive variant calls, resulting in inaccurate conclusions.

As shown below, a quality score of 20 represents an error rate of 1 in 100, with a corresponding call accuracy of 99%.

SBS Technology Overview

Illumina technology enables massively parallel sequencing with optimized SBS chemistry.

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Relationship Between Sequencing Quality Score and Base Call Accuracy
Quality Score Probability of Incorrect Base Call Inferred Base Call Accuracy
10 (Q10) 1 in 10 90%
20 (Q20) 1 in 100 99%
30 (Q30) 1 in 1000 99.9%
Flexibility for Multiple Sequencing Applications

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Illumina sequencing chemistry delivers high accuracy, with a vast majority of bases scoring Q30 and above. This level of accuracy is ideal for a range of sequencing applications, including clinical research.

For more in-depth information about sequencing quality scores, read the following technical notes:

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  1. Ewing B, Hillier L, Wendl MC, Green P. (1998): Base-calling of automated sequencer traces using phred. I. Accuracy assessment. Genome Res. 8(3):175-185
  2. Ewing B, Green P. (1998): Base-calling of automated sequencer traces using phred. II. Error probabilities. Genome Res. 8(3):186-194