Resource Files

Distributed as DRAGEN secondary analysis Product Files

It is recommended that reference files are based on normal (non-tumorous) samples processed in the same methods used for clinical samples, matching library prep, sequencer instrumentation, etc. When that is not possible, product files to support the analysis of samples prepared with the Illumina FFPE DNA Prep with Exome 2.5 Enrichment are available mostly on the DRAGEN Resources page and detailed here. Generally, CNV and MSI resource files show good performance across sequencing instruments, while systematic noise files are highly dependent on the sequence instrument and therefore important to match with clinical samples.

Resource	Prebuilt	Custom (Build your own)
Target BED	If no spike-in probes were included in enrichment, download from Illumina DNA Prep with Exome 2.5 BED files for hg19 or hg38 reference genomes.	If spike-in probes were included in enrichment, use a BED file of the combined coverage areas, ie., Exome 2.5 Plus Panel plus the custom panel. The requirements of a BED file are defined here.
Systematic noise files: considered essential for reducing false positive calls in Tumor-Only workflows, and they are also effective at improving precision in Tumor-Normal workflows.	DRAGEN Resources contains prebuilt systematic noise BED files. The directory SNV Somatic Systematic Noise v2.0.0 contains WES_*_v2.0.0_systematic_noise.snv.bed.gz noise files, which are built from a mixture of FF (fresh-frozen) and FFPE samples, with a mix of TruSeq PCR prep and Nextera prep and sequenced on the NovaSeq 6000. Reference genome builds (the * in the file name) include hg19 and hg38. What is downloaded from the Resource Page is a tarred directory. Extract the tarball, and within the untarred directory, find the systematic noise file. The g-zipped file can be uploaded to BSSH/ICA or used in the command-line execution of DRAGEN.	For instructions on how to build your own systematic noise file using internally-sequenced normal samples, see Custom Systematic Noise Files. Building your own noise file will be necessary if using spike-in probes. The normal samples used to generate the systematic noise file should match as closely as possible the sequencer, sample type, and library prep of the tumor samples. Also available are the DRAGEN Baseline Builder App on BSSH and the Systematic Noise File Builder on ICA.
CNV - Somatic pipelines: A Panel of normals (PONs) is used for calling gene amplification in tumor samples	DRAGEN Resources contains CNV Panel of Normals for Twist Bioscience for Illumina Exome FFPE 2.5 - DRAGEN 4.4 v1.0. The PON was generated from 45 FFPE benign adjacent samples from different tissue types and from male and female donors with the Illumina FFPE DNA Prep with Exome 2.5 Enrichment protocol. Libraries were sequenced on the NovaSeq 6000 sequencing system. Current supported builds include hg19 and hg38, both target.counts and gc-corrected.target.counts.	As with the systematic noise file, internally-sequenced normal samples can be used to generate a PON. The input should be the UMI-collapsed BAM files of the normal samples, and the reference genome and target BED need to match those used throughout the workflow. Also available are the DRAGEN Baseline Builder App on BSSH and the DRAGEN CNV Baseline Builder Pipeline on ICA.
MSI - Resource directory To be used when enabling the biomarker MSI in Tumor-only mode.	DRAGEN Resources contains Microsatellite Files v1.1.1. Within the WES directory are *.MSI_baselines_v1.1.0.combined.dist and *microsatellites.list files. The normal samples combined into this baseline files are from benign adjacent samples from different tissue types and from male and female donors. The normal samples were prepared with the Illumina FFPE DNA Prep with Exome 2.5 Enrichment protocol. Libraries were sequenced on the NovaSeq 6000 sequencing system. Current supported builds include hg19 and hg38. The list of microsatellite sites from which to calculate instabilityI are included in the *microsatellites.list files. Note that these microsatellites files do not contain sites on chrY, and that the same genome reference build should be used for the baseline combined dist and microsatellite list files. What is downloaded from the Resource Page is a tarred directory. Extract the tarball, and within the untarred directory, find the WES directory. Within that directory, are *.combined.dist based on different reference genome builds. Combined .dist files are input for DRAGEN v4.4.	See the DRAGEN manual for instructions on how to generate your own reference directory from interally-sequenced normal samples. Also available are the DRAGEN Baseline Builder App on BSSH and the DRAGEN MSI Baseline Builder 4-3-6 Pipeline on ICA. See the DRAGEN manual for how to generate custom microsatellite lists.