How to interpret clusters passing filter in run metrics
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Determining the number of reads passing filter (READS PF) is critical for evaluating the overall success of a sequencing run. Here are the step-by-step instructions for determining the number of clusters passing filter for a single lane, a single read, and a run using BaseSpace Sequence Hub (BSSH) or Sequencing Analysis Viewer (SAV).
How to access READS PF in the METRICS tab
Select a run.
Navigate to the METRICS tab.
Figure 1. Metrics tab in BSSH.
Locate the Per Read Metrics table.
Locate the READS PF column.
Values are the calculated number of READS PF per lane.
For a paired-end run, multiply this number by two to calculate the total number of READS PF per lane.
To obtain the number of READS PF for the entire run, add the total number of READS PF for each lane.
Figure 2. READS PF values are the calculated number of READS PF per lane. For a paired-end run, multiply this number by two to calculate the total number of READS PF per lane. For example, the NovaSeq run shown here has 2,313125,376 READS PF in Read 1 in lane 1. Because this is a paired-end run, the total number of READS PF in lane 1 is 4,626,250,752.
How to access READS PF in the INDEXING QC tab
Navigate to the INDEXING QC tab.
Locate the PF READS column per lane.
To switch between lanes, use the drop-down menu.
To obtain the number of READS PF for the entire run, add the total number of PF READS for each lane.
Note: The INDEXING QC metrics are calculated after demultiplexing, and therefore provide a more accurate read count than the metrics tab.
Figure 3. In the INDEXING QC tab, total number of reads passing filter are shown per lane. To switch between lanes, use the drop-down menu.
Navigate to the Summary tab.
The Cluster Count PF (M) column shows the number of reads passing filter in millions per lane and per read.
To obtain the total number of reads passing filter per read, add the values for a particular read. In this case, Read 1 contains 4,659,600,000 clusters passing filter.
Note: Unlike BaseSpace Sequence Hub, Sequencing Analysis Viewer shows metrics by read, not by lane.
Figure 4. In the Summary tab, the Cluster Count PF (M) column shows the number of reads passing filter in millions per lane and read. For example, Read 1 of lane 1 contains 2,313,130,000 reads passing filter. To obtain the total number of reads passing filter per read, add values 2,313,130,000 and 2,346,470,000. In this case, Read 1 contains 4,659,600,000 reads passing filter. The total number of reads passing filter for this run is ~9,319,200,000.
To determine the total number of reads passing filter for a run, add all the values in the Cluster Count PF column, excluding non-index reads.
The precise value is in the Indexing tab under the PF Reads column.
Figure 5. Selection pane on SAV for Indexes Passing Filter
Note: Values are represented slightly differently in BSSH and SAV. In both programs, the PF Reads tabs represent identical values, as do the Reads PF and Cluster Count PF (M) in the summary tabs.
For more information about Sequencing Analysis Viewer or BaseSpace Sequence Hub, visit their respective support pages:
For any feedback or questions regarding this article (Illumina Knowledge Article #6529), contact Illumina Technical Support techsupport@illumina.com.
