# Germline CNV Calling ASCN (WGS) The Germline WGS CNV caller (with Allele-Specific CN support) leverages depth of coverage and B-allele frequencies (BAFs) to detect germline copy number aberrations and regions with absence of heterozygosity (AOH). The caller requires in input the sample to analyze and a catalog of population SNPs (used to estimate the B-allele frequencies across the genome). The output file is a VCF file. Several components of other germline CNV callers are reused, such as [target](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/additional-documentation/cnv-preprocessing#target-counts) and [b-allele](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/additional-documentation/cnv-preprocessing#b-allele-counts-ascn-callers) counts estimation, [bias](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/additional-documentation/cnv-preprocessing#gc-bias-correction) correction, [normalization](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/additional-documentation/cnv-preprocessing#normalization) and [segmentation](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/cnv-calling/additional-documentation/cnv-segmentation). More details on the sample model selection are available in the [ASCN calling](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/cnv-calling/additional-documentation/cnv-ascn) section. See [Output Files](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/additional-documentation/cnv-output#output-files) for more details on output files. ![](https://3555446211-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2F-MflHs6hPlQAu-m5s2h_%2Fuploads%2Fgit-blob-dbfa7c4330a25cc90c8f2ba0ced4ed887443551f%2Fgermline-cnv-calling.WorkflowBlockDiagram.ASCN.png?alt=media) When enabled (default true), the caller can output an additional VCF with calls at resolutions compatible with cytogenetics technologies (e.g., ≥ 1 Mb, or whole-arm/whole-chromosome calls) For more information, see [Cytogenetics modality](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/cnv-calling/additional-documentation/cytogenetics-modality). ## Germline WGS CNV Calling (ASCN) Options You can use the following command-line options: | Option | Description | | ------------------------------------------------------------- | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | | `--fastq-file1`,`--fastq-file2`,`--bam-input`, `--cram-input` | Specify an input file. | | `--cnv-population-b-allele-vcf` | Specify a population SNP catalog. For more information on specifying b-allele loci, see [Specification of B-Allele Loci](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/additional-documentation/cnv-preprocessing#b-allele-counts-ascn-callers). | | `--enable-cnv` | Enable or disable CNV processing. Set to true to enable CNV processing. | | `--enable-map-align` | Enables the mapper and aligner module. | | `--output-directory` | Output directory where all results are stored. | | `--output-file-prefix` | Output file prefix that will be prepended to all result file names. | | `--ref-dir` | The DRAGEN reference genome hashtable directory. | | `--sample-sex` | If known, specify the sex of the sample. If the sample sex is not specified, the caller attempts to estimate the sample sex from alignments. | | `--cnv-enable-mosaic-calling` | Enable MOSAIC-calling mode (default true). For more information, see [MOSAIC-Calling Mode](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/additional-documentation/cnv-ascn#subclonalmosaic-calling-mode). | | `--cnv-enable-cyto-output` | Enable Cytogenetics-compatible output (default true). For more information, see [Cytogenetics Modality](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/cnv-calling/additional-documentation/cytogenetics-modality). | | `--cnv-enable-legacy-vcf-format` | Use VCF v4.2 for CNV VCF output (default false). | | `--cnv-enable-self-normalization` | Use self-normalization to normalize input sample. Default true. | ## Example command lines The following is an example command line: ``` dragen \ -r \ --output-directory \ --output-file-prefix \ --enable-map-align false \ --enable-cnv true \ --bam-input \ --cnv-enable-self-normalization true \ --cnv-population-b-allele-vcf \ --sample-sex ``` ## Differences with respect to Germline WGS CNV (Depth-only) The Germline WGS ASCN caller differs from the [depth-only caller](https://help.dragen.illumina.com/dragen-v4.4/product-guide/dragen-v4.4/dragen-dna-pipeline/cnv-calling/available-pipelines/germline-cnv-calling) in various respects: * Whereas both the depth-only caller and the ASCN caller make use of a depth of coverage profile, the ASCN caller (but not the depth-only caller) additionally makes use of allelic read counts at sites along the genome provided through a population SNP catalog. * The ASCN caller segments both profiles (depth and BAF), then integrates their results. The depth-only caller only segments the depth profile. * The ASCN caller estimates the most likely diploid coverage for the input sample using both profiles (depth and BAF), while the depth-only caller has only the depth profile at its disposal. * The ASCN caller can also detect AOH/LOH regions and mosaic alterations, while the depth-only caller is limited to DEL and DUP regions. Briefly, the following table summarizes the above differences: | Feature | Depth-only caller | ASCN caller | | ------------------------------------------ | ----------------- | ---------------------------------- | | Coverage Depth Profile | ✓ | ✓ | | B-Allele Frequency Profile | | ✓ | | Coverage Depth Segmentation | ✓ | ✓ | | B-Allele Frequency Segmentation | | ✓ | | DEL/DUP Detection | ✓ | ✓ | | Whole-arm/-chromosome Aneuploidy Detection | | ✓ (Requires Cytogenetics Modality) | | Whole-Genome Trisomy Detection | | ✓ | | Minor-Allele Copy-Number Estimation | | ✓ | | AOH/LOH Detection | | ✓ | | Mosaic Alterations Detection | | ✓ | ## Limitations The following features available in the non-ASCN germline caller are not yet supported: * Pedigree mode * Exome support * Star-Allele caller integration ## Examples The following are some examples of calls that can be output by the germline WGS ASCN caller: ``` # REF call chr1 819841 DRAGEN:REF:chr1:819841-4063702 N . 1000 PASS END=4063702;REFLEN=3243862 GT:CN:MCN:CNQ:MCNQ:CNF:MCNF:SM:SD:MAF:BC:AS:PE:OBF 0/0:2:1:1000:1000:2.00155:1.000775:1.000775:129.1:0.5:2661:6145:66,5:0.0037637 # Copy-neutral LOH call chr1 6104347 DRAGEN:CNLOH:chr1:6104348-6727324 N 1000 PASS END=6727324;REFLEN=622977;SVLEN=622977;LOHTYPE=AOH;SVTYPE=CNV GT:CN:MCN:CNQ:MCNQ:CNF:MCNF:SM:SD:MAF:BC:AS:PE:OBF 1/1:2:0:1000:1000:1.9876:0.001988:0.993798:128.2:0.001:528:916:10,12:0.00766703 # Example GAIN call without minor-allele information chr1 16605768 DRAGEN:GAIN:chr1:16605769-16645359 N 427 PASS END=16645359;REFLEN=39591;SVLEN=39591;SVTYPE=CNV GT:CN:MCN:CNQ:MCNQ:CNF:MCNF:SM:SD:MAF:BC:AS:PE ./1:6:.:1:.:6.27065:.:3.135326:404.457:.:23:0:6,11 # Example GAIN call with minor-allele information chr1 16715826 DRAGEN:GAIN:chr1:16715827-16949283 N 744 PASS END=16949283;REFLEN=233457;SVLEN=233457;SVTYPE=CNV GT:CN:MCN:CNQ:MCNQ:CNF:MCNF:SM:SD:MAF:BC:AS:PE:OBF 0/1:3:1:1000:99:3.08217:1.134239:1.541085:198.8:0.368:49:26:20,14:0.0384615 # Example GAIN LOH call chr15 20212550 DRAGEN:GAINLOH:chr15:20212551-20421468 N 390 PASS END=20421468;REFLEN=208918;SVLEN=208918;LOHTYPE=AOH;SVTYPE=CNV GT:CN:MCN:CNQ:MCNQ:CNF:MCNF:SM:SD:MAF:BC:AS:PE:OBF 1/1:6:0:1:1:5.90559:0.000000:2.952793:380.91:0:76:1:9,8:0 # Example MOSAIC GAIN call chr2 89673674 DRAGEN:GAIN:chr2:89673675-89851643 N 1000 PASS END=89851643;REFLEN=177969;MOSAIC;SVLEN=177969;SVTYPE=CNV GT:CN:MCN:CNQ:MCNQ:CNF:MCNF:SM:SD:MAF:BC:AS:PE ./1:4:.:0:.:4.32403:.:2.162016:278.9:.:70:0:13,2 # Example LOSS call chr1 25274774 DRAGEN:LOSS:chr1:25274775-25331683 N 226 PASS END=25331683;REFLEN=56909;SVLEN=56909;SVTYPE=CNV GT:CN:MCN:CNQ:MCNQ:CNF:MCNF:SM:SD:MAF:BC:AS:PE:OBF 0/1:1:0:1000:1000:1.01085:0.000000:0.505426:65.2:0:7:10:5,1:0 # Example MOSAIC LOSS call chrX 90207750 DRAGEN:LOSS:chrX:90207751-90458297 N 82 PASS END=90458297;REFLEN=250547;MOSAIC;SVLEN=250547;SVTYPE=CNV GT:CN:MCN:CNQ:MCNQ:CNF:MCNF:SM:SD:MAF:BC:AS:PE:OBF 0/1:1:0:0:0:1.6186:0.459684:0.809302:104.4:0.284:10:27:3,11:0 ```