CNV/SV calling analysis

VarSome Clinical offers two CNV calling solutions for your samples that are suitable for all types of NGS samples: WGS, WES and panels.

CNV calling analysis (from FASTQ)

VarSome Clinical currently offers two types of CNV calling solutions:

  • delly - suitable for single WGS samples and WGS tumor-normal CNV analysis.
  • exomeDepth - suitable for cohorts of WES/panels and also for WGS samples.

To start a CNV analysis from FASTQ, please go to "Launch analysis" > "CNV analysis from FASTQ".

Whole exome sequencing (WES) or targeted panel data

For such samples, we use the ExomeDepth CNV caller. The read depth based tool requires five or more (ideally between five and ten) germline or somatic samples that have already been analyzed on VarSome Clinical. These will be run as a cohort with each sample analyzed using a selection of the remaining samples as controls. The samples should all have been sequenced using the same assay since CNV calls will only be made within the assay's target regions. For optimal results, all samples should:

  1. be from the same sequencing run
  2. come from individuals unrelated to each other, and
  3. be of the same sex (either all male or all female). If the samples of the cohort are of not of the same sex, the CNV calls obtained for chromosomes X and Y will not be reliable.

All samples are analyzed together and the results (along with a visual display) of each sample are shown as a sub-analysis of that sample. 

Please note that an inherent limitation of WES is that it produces reads only covering the ~2% of the human genome that falls in exons. Therefore, the full spectrum of CNVs and breakpoints may not be completely characterized. In addition, many large CNVs and cross-chromosome events may not be detected. For optimal results, we suggest either sequencing the entire genome (WGS), or a different experimental approach such as array CGH. Nevertheless, CNV detection based on WES data may give an insight into CNV patterns for a specific disease or phenotype. For more details on the limitations of calling CNVs in such data, please see [1].

Sensitive mode

CNV calling for non-WGS CNV analyses is also available in “Sensitive mode”. Compared to standard mode, a lower CNV detection threshold is applied, resulting in more sensitive calling and typically in a higher number of calls. CNV detection can be particularly challenging; for instance single exon CNVs can be hard to call. Still, in a clinical setting, the ability to detect such CNVs is of paramount importance. Sensitive mode is optimised for the needs of clinical laboratories. It allows a shift to the trade-off between recall and false discovery to benefit sensitivity, compared to the standard mode.

You can enable this feature for either somatic or germline samples (or both) in Preferences.

Please, note that these settings are only available to the group administrator and any changes will be applied to all users of the group.

Whole genome sequencing (WGS) data

For WGS samples, VarSome Clinical offers two solutions:

  1. CNV calling for a single WGS sample. We use delly, an integrated structural variant (SV) caller tool that can detect both CNVs and other forms of Structural Variants (SVs) at single-nucleotide resolution in short-read genomic sequencing data. It combines 3 different approaches (paired-ends, split-reads and read-depth) to discover extensive genomic rearrangements. Quality passed CNV calls (deletions and duplications) are retained, while other types of SVs are currently not reported. 
  2. CNV calling for a cohort of (2-5) WGS samples. The ExomeDepth caller has been adapted to also process WGS. The solution is suitable for samples with long CNVs (>50kb) that may not be reliably called by Delly. For WGS, the assay target regions comprise the complete genome, split into 50Kb bins. As a result, this imposes a hard minimum size limit: no CNVs smaller than 50Kb can be detected using this approach. The requirements for non-relatedness between the samples and their processing by the same laboratory, sequencer and ideally in the same batch, apply to WGS samples too. All samples are analysed in a single CNV analysis and the results (along with a visual display) of each sample are shown as a sub-analysis.

A step-by-step example on how to run a CNV/SV analysis

Select “CNV analysis from fastq” from the “Launch analysis” drop-down menu on VarSome Clinical:

VarSome Clinical interface allows you to select a minimum of 5 and a maximum of 25 already analyzed samples to be used as a cohort for CNV calling. For best results, we recommend you select 5-10 samples from unrelated individuals of the same sex that were sequenced on the same sequencing run.

Each sample’s results will appear as a sub-analysis of the main analysis.

CNV annotation (from VCF)

Please see CNV annotation from VCF.

    See also


    [1] R. Tan et al., An Evaluation of Copy Number Variation Detection Tools from Whole-Exome Sequencing Data, Hum. Mut.,2014