Set Analysis Parameters
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1.
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Select the analysis name. |
The default name is the combination of the App name “UMI Error Correction”, and the current date (MM/DD/YYYY format) and time (HH:MM:SS format).
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2.
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Select Save Results To. |
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3.
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In the Select Projects drop-down menu, do one of the following. |
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Select an existing project to store the App results. |
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Select New to create a project. |
- Add a name and a description.
- Select Create to add the project to the projects list.
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5.
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In the Select Biosamples(s) drop-down menu, select up to 48 samples to be analyzed. |
Selected samples must be paired end reads (required). The sample list can be filtered using the Project filter on the right side of the sample selection interface.
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7.
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In the Panel Selection drop-down menu, select one of the following. |
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TruSight Tumor 170 DNA (manifest is preloaded) indicating that the TruSight Tumor 170 DNA panel was used to prepare the samples. |
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Custom (select a manifest below) indicating that a custom panel was used to prepare the sample. |
NOTE
This option requires the prior upload of an ILMN manifest file.
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8.
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If TruSight Tumor 170 DNA panel is not used, select Custom Manifest. |
The UMI Error Correction BaseSpace Sequence Hub App accepts any manifest files that conform to the following formatting rules based on Nextera Rapid Capture manifests.
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Manifest chromosomes corresponding to hg19 (eg chromosome 1 = chr1 and not chromosome 1 = 1) |
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[Regions] section (required) |
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Amplicon Start or Start |
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Data type by column header (column, data type) |
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Unique alphanumeric names for each manifest region |
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Inclusive manifest regions |
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One based indexing for start and end positions |
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Custom Manifest regions containing single base overlaps are not supported. |
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DownStream Probe Length |
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Data type by column header (column, data type) |
Target Region, string
Upstream Probe Length, int
Downstream Probe Length, int
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Recommended headers: Manifest File version and Reference Genome |
Example TruSight Tumor 170 DNA Manifest
#Experiment
#Final DNA Probes 04/28/2016
[Header]
Manifest Version Release2HDNA_manifest_160428
ReferenceGenome Homo_sapiens\UCSC\hg19\Sequence\WholeGenomeFASTA
[Regions]
Name Chromosome Amplicon Start Amplicon End Target Region Upstream Probe Length Downstream Probe Length
chr1_iSNP_1_1 chr1 4367257 4367353 chr1_iSNP_1 0 0
PIK3CD_Exon3_2 chr1 9770514 9770654 PIK3CD_Exon3 0 0
PIK3CD_Exon4_3 chr1 9775599 9775827 PIK3CD_Exon4 0 0
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9.
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Configure Analysis Parameters. |
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Minimum Map Quality – (Default = 0) |
The minimum map quality parameter allows you to filter each read based on the quality of the alignment (MapQ value calculated using BWA). A MapQ value of 0 indicates a read with an ambiguous alignment as calculated by BWA. The minimum map quality default setting is 0 (to ensure maximum read utilization) and can be set up to 60. This filter is applied at the read collapsing step, which means the aligned BAM output will contain all reads.
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Minimum Supporting Reads – (Default = 2) |
The minimum supporting reads parameter allows you to specify the minimum number of supporting reads in a collapsed family in the final output of the collapsed BAM file. The default setting is 2 and can be set from 1 to 20. Collapsed read families with fewer than specified supporting reads are discarded. The number of reads that went into a given collapsed read can be found in the XV and XW tags of the reads in the collapsed BAM file. The specified minimum supporting reads is the sum of the two tags, where the XV tag specifies the number of read pairs used that were in the forward read orientation and the XW tag specifies any used with reverse read orientations.
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10.
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After all analysis parameters are set, select Start Analysis. |
The Analysis Info page is displayed, which shows the status of the analysis in the Status field. An email is sent to the registered email address after the analysis is completed.