# DNA Germline WGS UMI The DRAGEN recipe includes the recommended pipeline specific commands. A DRAGEN recipe is a predefined set of analysis parameters and workflow settings tailored for a specific type of genomic analysis. Some default parameters are included for clarity and are marked with comments. ``` /opt/dragen/$VERSION/bin/dragen #DRAGEN install path --ref-dir $REF_DIR #path to DRAGEN graph hashtable --output-directory $OUTPUT --intermediate-results-dir $PATH #e.g. SDD /staging --output-file-prefix $PREFIX # Inputs --fastq-list $PATH #see 'Input Options' for FQ, BAM or CRAM --fastq-list-sample-id $STRING # Mapper --enable-map-align true #optional with BAM/CRAM input --enable-map-align-output true #optionally save the output BAM --enable-sort true #default=true # UMI --umi-enable true --umi-source STRING #Default='qname' --umi-library-type STRING #e.g. random-duplex --umi-metrics-interval-file $BED --remove-duplicates false --umi-min-supporting-reads 1 #Default=2 # Small variant caller --enable-variant-caller true # Annotation --variant-annotation-data PATH --variant-annotation-assembly GRCh37/8 --enable-variant-annotation true # SV --enable-sv true # CNV --enable-cnv true --cnv-enable-self-normalization true # HLA genotyper --enable-hla true --hla-enable-class-2 true #optional if assay covers class II HLA regions # Targeted caller --enable-targeted true #Targeted # Star allele --enable-star-allele true # PGX --enable-pgx true #PGX # Short tandem repeats --repeat-genotype-enable true ``` ## Notes and additional options ### Hashtable For DRAGEN germline runs, it is recommended to use the graph hashtable. See: [Product Files](https://support.illumina.com/sequencing/sequencing_software/dragen-bio-it-platform/product_files.html) ### Input options DRAGEN input sources include: fastq list, fastq, bam, or cram. FQ list Input ``` --fastq-list $PATH --fastq-list-sample-id $STRING ``` FQ Input ``` --fastq-file1 $PATH --fastq-file2 $PATH --RGSM $STRING --RGID $STRING ``` BAM Input ``` --bam-input $PATH ``` CRAM Input ``` --cram-input $PATH ``` ### Mapping and Aligning | Option | Description | | -------------------------------- | ---------------------------------------------------------------------------------------------------- | | `--enable-map-align true` | Optionally disable map & align (default=true). | | `--enable-map-align-output true` | Optionally save the output BAM (default=false). | | `--Aligner.clip-pe-overhang 2` | Clean up any unwanted UMI indexes. Only use when reads contain UMIs, but UMI collapsing was not run. | ### UMI | Option | Description | | ---------------------------------- | ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | | `--umi-source STRING` | Specify the input type for the UMI sequence. Options: `qname`, `fastq`, `bamtag`. | | `--umi-library-type STRING` | Set the batch option for different UMIs correction. Options: `random-duplex`, `random-simplex`, `nonrandom-duplex`. | | `--umi-nonrandom-whitelist $PATH` | If UMI is nonrandom, either a whitelist or correction table is required. The whitelist includes a valid UMI sequence per line. | | `--umi-correction-table $PATH` | If UMI is nonrandom, either a whitelist or correction table is required. The correction table defaults to the table used by TruSight Oncology: \/resources/umi/umi\_correction\_table.txt.gz. | | `--umi-min-supporting-reads INT` | Specify the number of matching UMI input reads required to generate a consensus read. Any family with insufficient supporting reads is discarded. The default is 2, but most pipelines perform better with this setting set to 1. A setting of 2 may potentially be relevant for samples with ultra deep coverage (e.g. ctDNA). | | `--umi-metrics-interval-file $BED` | Target region in BED format. | | `--umi-emit-multiplicity both` | Set the consensus sequence type to output. DRAGEN UMI allows collapsing duplex sequences from the two strands of the original molecules. For more information, see [Merge Duplex UMIs](https://help.dragen.illumina.com/dragen-v4.3/product-guide/dragen-dna-pipeline/unique-molecular-identifiers#merge-duplex-umis). | | `--umi-start-mask-length INT` | Number of additional bases to ignore from start of read. The default is 0. To reduce FP optionally set to 1. | | `--umi-end-mask-length INT` | Number of additional bases to ignore from end of read. The default is 0. To reduce FP optionally set to 3. | For more information see: [UMI Options](https://help.dragen.illumina.com/dragen-v4.3/product-guide/dragen-dna-pipeline/unique-molecular-identifiers#umi-options). ### SNV DRAGEN SNV VC employs machine learning based variant recalibration (DRAGEN-ML). It processes read and other contextual evidence to remove false positives, recover false negatives and reduce zygosity errors. No additional setup is required. DRAGEN-ML is enabled by default as needed, when running the germline SNV VC on hg19 or hg38. Note that we do not recommend changing the default QUAL thresholds of 3 for DRAGEN-ML and 10 for DRAGEN without ML. These values differ from each other because DRAGEN-ML improves the calibration of QUAL scores, leading to a change in the scoring range. | Option | Description | | ------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------- | | `--vc-target-bed` | Limit variant calling to region of interest. | | `--vc-combine-phased-variants-distance INT` | Maximum distance over which phased variants will be combined. Set to 0 to disable. Valid range is \[0; 15] (Default=2) | | `--vc-emit-ref-confidence GVCF` | To enable gVCF output. | | `--vc-enable-vcf-output` | To enable VCF file output during a gVCF run, set to true. The default value is false. | For more detail on the small variant caller in somatic mode please refer to [Somatic Mode](https://help.dragen.illumina.com/dragen-v4.3/product-guide/dragen-v4.3/dragen-dna-pipeline/small-variant-calling/somatic-mode) ### Annotation For instructions on how to download the Nirvana annotation database, please refer to [Nirvana](https://help.dragen.illumina.com/dragen-v4.3/product-guide/dragen-v4.3/nirvana) ### HLA | Option | Description | | --------------------------------- | ------------------------------------------------------------------------------------------------------------------------------- | | `--enable-hla` | Enable HLA typer (this setting by default will only genotype class 1 genes) | | `--hla-as-filter-min-threshold` | Internal option to set min alignment score threshold. The default is 59 and works for WES and WGS. Set to 29 for panels. | | `--hla-as-filter-ratio-threshold` | Minimum Alignment score of a read mate to be considered. The default is 0.67 and works for WES and WES. Set to 0.85 for panels. | | `--hla-enable-class-2` | Extend genotyping to HLA class 2 genes (default=true). | ### CNV | Option | Description | | ------------------------------------- | --------------------------------------------------------------------------------------------------------------------------------------------------------- | | `--cnv-enable-gcbias-correction true` | Enable or disable GC bias correction when generating target counts. | | `--cnv-segmentation-mode $SEG_MODE` | Option to override the default segmentation algorithm. Defaults include `slm` for germline WGS, `aslm` for somatic WGS, and `hslm` for targeted analysis. | For more information, see [CNV Calling](https://help.dragen.illumina.com/dragen-v4.3/product-guide/dragen-v4.3/dragen-dna-pipeline/cnv-calling).