2023 ACMG Annual Clinical Genetics Meeting Digital Edition: Poster Gallery-ROH Pipeline Enables Analysis of Regions of Homozygosity Using Next Generation Sequencing Data

Pdf Summary

The document describes a pipeline developed for the analysis of regions of homozygosity (ROH) in Prader-Willi Syndrome (PWS) using next-generation sequencing (NGS) data. ROH are long stretches of homozygous DNA resulting from uniparental isodisomy or autozygosity, and they can be associated with disease when they overlap with deleterious variants or imprinted regions. PWS is a rare genetic disorder caused by the loss of paternally inherited genes on chromosome 15.<br /><br />The ROH pipeline utilizes Automap to call ROH for each sample and performs post-processing steps such as removing gap regions, combining ROH calls into blocks, filtering based on size, and determining false positive calls using coverage data. In addition, the pipeline uses AnnotSV for annotation of filtered ROH blocks and knotAnnotSV to generate interactive visualizations for interpretation based on phenotype associations.<br /><br />Verification of reported ROH is essential, achieved through visualization with B-allele plots and sequence reads alignment. The pipeline was applied to analyze 49 PWS samples, accurately identifying cases of chromosome 15 segmental isodisomy and ruling out heterozygous deletions mimicking ROH.<br /><br />The pipeline was implemented using Snakemake for workflow management, Python for scripting, and Conda environments within a Singularity container for portability. The pipeline supports input of VCF files, sample names, and phenotypes coded with Human Phenotype ontology (HPO) IDs.<br /><br />The document includes figures illustrating the tasks performed by the ROH pipeline and examples of raw and filtered ROH in chromosome 15 of PWS patient samples.<br /><br />In summary, the ROH pipeline presented in this document streamlines the analysis of ROH in PWS using NGS data, providing efficient calling, filtering, annotation, and verification steps. It is useful for identifying cases of uniparental isodisomy in chromosome 15 and differentiating them from other genetic variants.

Asset Subtitle

Presenting Author - Manavalan Gajapathy, PhD; Co-Author - Brandon M. Wilk, BS; Co-Author - Donna M. Brown, MS; Co-Author - Caroline Vrana-Diaz, PhD; Co-Author - Jessica Bohonowych, PhD; Co-Author - Theresa Strong, PhD; Co-Author - Elizabeth A. Worthey, PhD;

Meta Tag

Bioinformatics

Exome sequencing

Genome sequencing

Genomic Methodologies

Imprinting

Methodology

Uniparental Disomy

Co-Author Brandon M. Wilk, BS

Co-Author Donna M. Brown, MS

Co-Author Caroline Vrana-Diaz, PhD

Co-Author Jessica Bohonowych, PhD

Co-Author Theresa Strong, PhD

Co-Author Elizabeth A. Worthey, PhD

Presenting Author Manavalan Gajapathy, PhD

Keywords

pipeline

regions of homozygosity

ROH

Prader-Willi Syndrome

PWS

next-generation sequencing

NGS data

Automap

AnnotSV

knotAnnotSV