Analyze QIAseq Hybrid Capture DNA in Trio
The Analyze QIAseq Hybrid Capture DNA in Trio (Illumina) template workflow can be used for analyzing DNA reads generated using the QIAseq Multimodal DNA Library Kit (without UMIs) or QIAseq FX DNA Library Kit followed by hybrid capture-based target enrichment, such as QIAseq Exome or QIAseq xHYB Human.
The workflow identifies putative disease causing, inherited variants in a family of three, where there is an affected parent, an unaffected parent, and a proband.
The workflow includes all necessary steps for processing and analyzing the reads:
- Target coverage statistics are generated using QC for Targeted Sequencing.
- Reads are trimmed using Trim Reads.
- Reads are mapped to the human reference genome using Map Reads to Reference.
- A guidance track is generated from the mapped reads using Structural Variant Caller.
- An improved mapping is obtained by realigning the mapped reads using the guidance track and Local Realignment.
- Variants are detected in the relevant target regions from the improved mapping using Fixed Ploidy Variant Detection.
- The variants are annotated with various information, such as overlap with repeat/homopolymer regions and genes, and are subsequently filtered to remove likely artifacts using Filter on Custom Criteria.
- The putative disease causing variants are identified as the variants present in both affected individuals but not in the unaffected individual.
- A combined summary report is created using Combine Reports.
Launching the workflow
To run this workflow, go to:
Workflows | Template Workflows | Biomedical Workflows (
) | QIAseq Sample Analysis (
) | QIAseq DNA workflows (
) | Analyze QIAseq Hybrid Capture in Trio (
)
See Launching workflows individually and in batches for general information.
Options can be configured in the following wizard steps:
- Choose where to run. If you are connected to a CLC Server via the CLC Workbench, you will be asked where you would like to run the analysis. We recommend that you run the analysis on a CLC Server when possible.
- Select Reads from proband/affected parent/unaffected parent. Select the input reads from proband, affected parent, and unaffected parent, respectively. When analyzing more than one trio at a time, check Batch in the lower left corner of the dialog.
- Specify reference data handling. Select the QIAseq DNA Hybrid Capture and WGS hg38 Reference Data Set, see Reference data management for details.
- Configure batching, if running the workflow in batch mode. Define the batch units.
- Batch overview, if running the workflow in batch mode. Verify that the batching is as intended.
- Target regions. Choose the relevant target regions from the drop down list. If the data was produced using a panel that is not available in the list, see QIAseq custom panels.
- Map Reads to Reference. Configure masking. By default, regions defined by the Genome Reference Consortium are excluded to remove false duplications, including one involving U2AF1.
This can be configured separately for proband, affected parent, and unaffected parent. The same options should be applied to all three.
- Fixed Ploidy Variant Detection. Configure the Minimum frequency (%) as needed, if variants should be detected at lower frequencies. Note that the value should be no more than half of the frequency specified when filtering variants.
This can be configured separately for proband, affected parent, and unaffected parent. The same options should be applied to all three.
- Filter on Custom Criteria (proband). Configure how variants should be filtered. The default options are optimized for samples with relatively high quality and coverage providing a strong balance between sensitivity and precision. For lower-quality or lower-coverage samples, when seeking a different sensitivity-precision balance, or for detecting low-frequency variants, additional filters or adjusted thresholds might be needed. See Filter on Custom Criteria for details on how to configure the options.
- Result handling. Choose if a workflow result metadata and/or log should be saved.
- Save location for new elements. Choose where to save the data, and press Finish to start the analysis.
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