Analyze QIAseq Methyl
The Analyze QIAseq Methyl (Illumina) template workflow can be used to call methylation levels on data generated with QIAseq Targeted Methyl Panels.
The workflow includes all necessary steps for processing and analyzing the reads:
- Various statistics summarizing and visualizing the input reads are produced using QC for Sequencing Reads.
- UMIs are removed using Remove and Annotate with Unique Molecular Index.
- Reads are trimmed using Trim Reads.
- Reads are mapped using Map Bisulfite Reads to Reference.
- UMI reads are created using Calculate Unique Molecular Index Groups and Create UMI Reads from Grouped Reads.
- Target coverage statistics are generated using QC for Targeted Sequencing.
- Differential methylation is detected using Call Methylation Levels.
- Sample composition is optionally predicted using Predict Methylation Profile.
- A summary report is created using Create Sample Report.
Launching the workflow
To run this workflow, go to:
Workflows | Template Workflows | Biomedical Workflows (
) | QIAseq Sample Analysis (
) | Other QIAseq Workflows (
) | Analyze QIAseq Methyl (Illumina) (
)
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.
- Specify workflow path. Select whether methylation profile prediction should be performed. Only select 'Yes' for data produced with T Cell Infiltration panels such as the 'T Cell Infiltration (MHS-202Z)' panel.
- Select Reads. Select the input reads. When analyzing more than one sample at a time, check Batch in the lower left corner of the dialog.
- Specify reference data handling.
Select the QIAseq Methyl Panels hg19 Reference Data Set, see Reference data management for details.
Note that the QIAseq Targeted Methyl Panels are designed to use hg19 as reference, but custom panels can be designed against either hg19 or hg38. When analyzing data from a custom panel designed against hg38, a Custom Reference Data Set must first be created. See also Import QIAGEN Primers.
- 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 primers. Choose the relevant primers from the drop down list.
Specify the primers, either by selecting them from the Reference Data Set (QIAseq Methyl Panels hg19) for QIAGEN panels or, for custom panels, by importing them as described in Import QIAGEN Primers
- 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 Bisulfite Reads to Reference. Specify whether a 'Directional' (reads from both strands) or 'Non-directional' protocol was used to generate the data.
- QC for Targeted Sequencing. Specify the minimum coverage needed on all positions in a target for it to be considered covered.
- Predict Methylation Profile, if predicting methylation profile. Specify the minimum coverage a CpG site must have to be used in the prediction.
- Create Sample Report. Select relevant summary items and configure thresholds for quality control. These are included in the quality control section of the sample report. The default values are appropriate for most data sets, but may need to be adjusted.
- 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.
Launching using the QIAseq Panel Analysis Assistant
The workflow is also available in the QIAseq Panel Analysis Assistant under Targeted Methyl.
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