DRAGEN v4.3.17 Recipe

Detailed instructions on how to configure the DRAGEN command-line for analyzing samples processed with the Illumina FFPE DNA Prep with Exome 2.5 Enrichment.

The following DRAGEN recipes are specific for Illumina FFPE DNA Prep with Exome 2.5 Enrichment libraries. A general recipe for Somatic analysis of Tumor Normal samples with UMI is provided on DRAGEN v4.4 support pages.

/opt/dragen/$VERSION/bin/dragen        #DRAGEN install path 
--ref-dir $REF_DIR                     #path to DRAGEN linear hashtable 
--output-directory $OUTPUT 
--intermediate-results-dir /staging    #tmp dir on fast SDD 
--output-file-prefix $PREFIX 
# Inputs
--tumor-fastq-list $PATH                #see 'Input Options' for FQ, BAM or CRAM 
--tumor-fastq-list-sample-id $STRING 
--fastq-list $PATH                      #see 'Input Options' for FQ, BAM or CRAM 
--fastq-list-sample-id $STRING 
# Mapper
--enable-map-align true
--enable-map-align-output true         #save the output BAM (default=false)
--qc-coverage-ignore-overlaps true     #do not double-count overlapping mates
# UMI
--umi-enable true 
--umi-library-type nonrandom-duplex
--umi-metrics-interval-file $VC_TARGET_BED #see Resource Files page
--umi-min-supporting-reads 1
--umi-start-mask-length 1
--umi-end-mask-length 3
# Small variant caller 
--enable-variant-caller true 
--vc-target-bed $VC_TARGET_BED         #see Resource Files page
--vc-systematic-noise $PATH            #see Resource Files page
--vc-enable-umi-solid true 
--vc-sq-call-threshold 3 
--vc-sq-filter-threshold 15         
# CNV 
--enable-cnv true
--cnv-use-somatic-vc-baf true
--cnv-target-bed $VC_TARGET_BED 
--cnv-combined-counts $PATH            #see Resource Files page
# Annotation                           #annotation is required if enabling TMB
--enable-variant-annotation true
--variant-annotation-data $PATH        #see notes below
--variant-annotation-assembly GRCh37/8
# TMB 
--enable-tmb true
# HRD Scoring 
--enable-hrd true                       #requires CNV 
# Microsatellite Instability (MSI) 
--msi-command tumor-only
--max-base-quality 63                                    #default if UMI is enabled
--msi-coverage-threshold 60                              #see notes below 
--msi-microsatellites-file ${microsatellite_file}        #see Resource Files page
--msi-ref-normal-dir ${normal_reference_directory}       #see Resource Files page

Notes and additional options

Hashtable

For DRAGEN somatic runs it is recommended to use the linear (non-graph) hashtable.

Please see: DRAGEN references

Inputs

Here are support pages for the formats of different input file types.

FQ list Input

--fastq-list $PATH 
--fastq-list-sample-id $STRING 

or

--tumor-fastq-list $PATH 
--tumor-fastq-list-sample-id $STRING 

FQ Input

--fastq-file1 $PATH 
--fastq-file2 $PATH 
--RGSM $STRING 
--RGID $STRING 

or

--tumor-fastq1 $PATH 
--tumor-fastq2 $PATH 
--RGSM-tumor $STRING 
--RGID-tumor $STRING 

BAM Input

--bam-input $PATH

or

--tumor-bam-input $PATH

CRAM Input

--cram-input $PATH

or

--tumor-bam-input $PATH

Mapper

Option	Description
--qc-coverage-ignore-overlaps true	resolve all of the alignments for each fragment and avoid double-counting any overlapping bases

UMI

The above recipe details UMI options specific for the Illumina FFPE DNA Prep with Exome 2.5 Enrichment Library kit.

SNV

The vc-sq-filter-thresholdflag can be used to fine-tune the "somatic quality" value at which variants are called as true somatic variants. Variants with values between the vc-sq-call-threshold and vc-sq-filter-threshold values are labeled with the Filter ID of weak_evidence.

Annotation

Top directory containing Nirvana data file. Instructions on how to download the resource are at https://support.illumina.com/content/dam/illumina-support/help/Illumina_DRAGEN_Bio_IT_Platform_v3_7_1000000141465/Content/SW/Informatics/Dragen/Nirvana_DownloadData_fDG.htm. DRAGEN is expecting a top directory containing the sub-directories Cache, References, Supplementary Annotation.

Microsatellite Instability (MSI)

Microsatellite Instability (MSI) is run in tumor-only mode, but other variants and biomarkers are calculated in tumor-normal mode. msi-coverage-threshold is a required parameter for both tumor-only and tumor-normal, and should be set according to the sample coverage. The sites in the msi-microsatellites-file need to match those in the dist files within the msi-ref-normal-dir .