Quality Scoring
A quality score, or Q-score, is a prediction of the probability of an incorrect base call. A higher Q-score
implies that a base call is higher quality and more likely to be correct.
The Q-score is a compact way to communicate small error probabilities. Q(X) represents quality scores,
where X is the score. The following table shows the relationship between the quality score and error
probability.
Q-score Q(X)
Error Probability
Q40
0.0001 (1 in 10,000)
Q30
0.001 (1 in 1,000)
Q20
0.01 (1 in 100)
Q10
0.1 (1 in 10)
NOTE
Quality scoring is based on a modified version of the Phred algorithm.
Quality scoring calculates a set of predictors for each base call, and then uses the predictor values to look up
the Q-score in a quality table. Quality tables are created to provide optimally accurate quality predictions for
runs generated by a specific configuration of sequencing platform and version of chemistry.
After the Q-score is determined, results are recorded in the base call files.
Q-Score Binning
RTA groups quality scores into specific ranges, or bins, and assigns a value to each range. Q-score binning
significantly reduces storage space requirements without affecting accuracy or performance of downstream
applications.
Q-score binning contributes to the efficiency of analysis processes and data transfer requirements associated
with the high throughput of the HiSeq 2000. The resulting *.bcl file is smaller because the compression
algorithms are able to compress the file more effectively. Less data are written to the instrument computer
and transferred to a network location, making the file copy faster.
Monitor Run Metrics
RTA automatically generates quality metrics when image analysis begins. However, not all metrics are
available at the early cycles because some processes require multiple cycles to generate data.
Data
Cycle
Image
analysis
After cycle 5.
During the first 5 cycles of the run, RTA generates a template of cluster locations.
Base
calls
After cycle 12.
Base calling begins after the color matrix is estimated at cycle 12.
Phasing
estimates
After cycle 25.
The phasing corrections for the first 25 cycles determine the phasing estimate.
Quality
scores
After cycle 25.
A quality score is generated for reads that pass the quality filter. Because quality scores require corrected
intensities from future cycles, quality scoring always follows base calling.
Document # 15011190 v03
For Research Use Only. Not for use in diagnostic procedures.
59
HiSeq 2000 System Guide