View Run Charts
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The Charts tab displays graphs of run metrics that update as the run progresses. Use these charts to monitor the quality of the run.
Data by Cycle graphs the progression by cycle of quality metrics during a run.
The Data by Cycle chart has the following features:
You can select the displayed metric, surface (if your system scans multiple surfaces), cycle, and channel (base) using the drop-down lists.
The chart is displayed with tailored scaling by default, or you can fix the Y-axis scale by selecting the Fix Scale checkbox.
Some metrics (% ≥Q20 and % ≥Q30) are monitored for a single cycle by default, or you can monitor the cumulative metrics (up to that cycle) by selecting the Accum checkbox.
You can focus on an area of interest by dragging to pan the view or using the mouse wheel to zoom in.
The following table details the possible metrics displayed in this chart. The available options vary by run.
Setting
Description
Intensity
This chart shows the intensity by color and cycle of the 90% percentile of the data for each tile.
FWHM
The percentage of clusters for which the selected base (A, C, T, or G) has been called.
% Base
The percentage of clusters for which the selected base (A, C, T, or G) has been called.
% NoCall
The percentage of clusters that have no call.
% ≥Q20 and % ≥Q30
The percentage of bases with a quality score of > 20 or > 30, respectively. These charts are generated after the 25th cycle, and the values represent the current scored cycle.
Median Q-Score
The median Q-score for each tile over all bases for the current cycle. These charts are generated after cycle 25. Use this setting to examine the Q-scores of your run as it progresses. Because it relies on a single threshold, the %≥Q30 plot can be oversimplified.
Error Rate
The calculated error rate, as determined by a spiked in PhiX control sample. If no PhiX control sample is run in the lane, this chart is not available.
Phasing, Prephasing
The estimated percentage of molecules in a cluster for which sequencing falls behind (phasing) or jumps ahead (prephasing) the current cycle within a read.
Legacy Phasing Rate, Legacy Prephasing Rate
The estimated percentage of molecules in a cluster for which sequencing falls behind (phasing) or jumps ahead (prephasing) the current cycle within a read. Legacy rates consider only the first 25 cycles.
Corrected Intensity
The intensity corrected for cross talk between the color channels by the matrix estimation and phasing and prephasing.
Called Intensity
The intensity for the called base.
Signal to Noise
The signal to noise ratio is calculated as mean called intensity divided by standard deviation of noncalled intensities.
Q-Score Distribution graphs the number of bases by quality score. The quality score is cumulative for the current cycle. Only bases from reads that pass the quality filter are included.
Use it to evaluate the Q-Score distribution for a run, which is an excellent indicator for run performance.
The Q-Score Distribution chart has the following features:
You can select the displayed read and cycle using the drop-down lists.
The chart is displayed with tailored scaling by default, or you can fix the Y-axis scale by selecting the Fix Scale checkbox.
Some metrics (% ≥Q20 and % ≥Q30) are monitored for a single cycle by default, or you can monitor the cumulative metrics (up to that cycle) by selecting the Accum checkbox.
You can focus on an area of interest by dragging to pan the view or using the mouse wheel to zoom in.
The Flow Cell Chart shows color-coded quality metrics per tile for the entire flow cell.
Use the Flow Cell Chart to judge local differences per cycle, per lane, or per read in sequencing metrics on a flow cell. It is also an easy way to see the %≥Q30 metric, which is an excellent single metric to evaluate a run. Do not use the Flow Cell Chart to look at downstream analysis metrics.
The Flow Cell chart has the following features:
You can select the displayed metric, surface (if your system scans multiple surfaces), cycle, and channel (base) using the drop-down lists.
The color bar to the right of the chart indicates the values that the colors represent.
The chart is displayed with tailored scaling by default, or you can fix the Y-axis scale by selecting the Fix Scale checkbox.
Some metrics (% ≥Q20 and % ≥Q30) are monitored for a single cycle by default, or you can monitor the cumulative metrics (up to that cycle) by selecting the Accum checkbox.
Tiles that have not been measured or are not monitored are gray.
You can focus on an area of interest by dragging to pan the view or using the mouse wheel to zoom in.
The following table details the possible metrics displayed in this chart. The available options vary by run.
Option
Description
Intensity
This chart shows the intensity by color and cycle of the 90% percentile of the data for each tile.
FWHM
The average full width of clusters at half maximum (in pixels). Used to display focus quality.
% Base
The percentage of clusters for which the selected base (A, C, T, or G) has been called.
% NoCall
The percentage of clusters that have no call.
% ≥Q20 and% ≥Q30
The percentage of bases with a quality score of > 20 or > 30, respectively. These charts are generated after the 25th cycle, and the values represent the current scored cycle.
Median Q-Score
The median Q-score for each tile over all bases for the current cycle. These charts are generated after cycle 25. Use this setting to examine the Q-scores of your run as it progresses. Because it relies on a single threshold, the %≥Q30 plot can be oversimplified.
Density
The density of clusters for each tile (in thousands per mm2).
Density PF
The density of clusters passing filter for each tile (in thousands per mm2).
Clusters
The number of clusters for each tile (in millions).
Clusters PF
The number of clusters passing filter for each tile (in millions).
Error Rate
The calculated error rate, as determined by a spiked in PhiX control sample. If no PhiX control sample is run in the lane, this chart is not available.
Phasing, Prephasing
The estimated percentage of molecules in a cluster for which sequencing falls behind (phasing) or jumps ahead (prephasing) the current cycle within a read.
Legacy Phasing Rate, Legacy Prephasing Rate
The estimated percentage of molecules in a cluster for which sequencing falls behind (phasing) or jumps ahead (prephasing) the current cycle within a read. Legacy rates consider only the first 25 cycles.
% Aligned
The percentage of reads from clusters in each tile that aligned to the PhiX genome.
Occupied Count (K)
The total number of wells (in thousands) on the flow cell containing clusters with DNA usable for sequencing. Wells with at least nine G bases called in the first 10 cycles are considered empty. Low occupancy rate combined with low %PF may indicate the loading concentration is too low. High occupancy rate combined with suboptimal %PF may indicate the loading concentration is too high.
% Occupied
The percentage of wells (for patterned flow cells) or nonduplicated spots (for nonpatterned flow cells) on the flow cell containing clusters. Wells with at least nine G bases called in the first 10 cycles are considered empty.
Corrected Intensity
The intensity corrected for cross talk between the color channels by the matrix estimation and phasing and prephasing.
Called Intensity
The intensity for the called base.
Signal to Noise
The signal to noise ratio is calculated as mean called intensity divided by standard deviation of noncalled intensities.
Note the variable scales used on these different parameters.
The Data by Lane chart plots allow you to view quality metrics per lane.
Use the Data By Lane plot to examine the difference in quality metrics between lanes. Do not use the Data By Lane plot to look at alignment or variant calling analysis metrics.
The Data by Lane plot has the following features:
You can select the metric, surface, and read from the drop-down lists.
The plots show the distribution of mean values for a given parameter across all tiles in a given lane.
The red line indicates the median tile value for the parameter displayed. Blue boxes are for raw clusters, green boxes for clusters passing filter.
The box outlines the interquartile range (the middle 50% of the data) for the tiles analyzed for the data point.
The error bars delineate the minimum and maximum without outliers.
The outliers are the values that are more than 1.5 times the interquartile range below the 25th percentile, or more than 1.5 times the interquartile range above the 75th percentile. Outliers are indicated as dots.
You can focus on an area of interest by dragging to pan the view or using the mouse wheel to zoom in.
The following table details the possible metrics displayed in this plot. The available options vary by run.
Setting
Description
Density
The density of clusters for each tile (in thousands per mm2).
Clusters
The number of clusters for each tile (in millions).
Phasing, Prephasing
The estimated percentage of molecules in a cluster for which sequencing falls behind (phasing) or jumps ahead (prephasing) the current cycle within a read.
Legacy Phasing Rate, Legacy Prephasing Rate
The estimated percentage of molecules in a cluster for which sequencing falls behind (phasing) or jumps ahead (prephasing) the current cycle within a read. Legacy rates consider only the first 25 cycles.
% Aligned
The percentage of reads from clusters in each tile that aligned to the PhiX genome.
Occupied Count (K)
The total number of wells (in thousands) on the flow cell containing clusters with DNA usable for sequencing. Wells with at least nine G bases called in the first 10 cycles are considered empty. Low occupancy rate combined with low %PF may indicate the loading concentration is too low. High occupancy rate combined with suboptimal %PF may indicate the loading concentration is too high.
% Percent Occupied
The percentage of wells (for patterned flow cells) or nonduplicated spots (for nonpatterned flow cells) on the flow cell containing clusters. Wells with at least nine G bases called in the first 10 cycles are considered empty.
% PF
The percentage of clusters passing filter that are assigned to the index.
The Q-Score Heatmap provides an overview of quality scores across cycles.
The Q-Score Heatmap has the following features:
The vertical color bar indicates the value that each color represents.
Scaling is specific to the run, representing 0%–100% of the maximum value.
