Dataset Information

Linked-read sequencing for detecting short tandem repeat expansions.

ABSTRACT: Detection of short tandem repeat (STR) expansions with standard short-read sequencing is challenging due to the difficulty in mapping multicopy repeat sequences. In this study, we explored how the long-range sequence information of barcode linked-read sequencing (BLRS) can be leveraged to improve repeat-read detection. We also devised a novel algorithm using BLRS barcodes for distance estimation and evaluated its application for STR genotyping. Both approaches were designed for genotyping large expansions (> 1 kb) that cannot be sized accurately by existing methods. Using simulated and experimental data of genomes with STR expansions from multiple BLRS platforms, we validated the utility of barcode and phasing information in attaining better STR genotypes compared to standard short-read sequencing. Although the coverage bias of extremely GC-rich STRs is an important limitation of BLRS, BLRS is an effective strategy for genotyping many other STR loci.

SUBMITTER: Chiu R

PROVIDER: S-EPMC9174224 | biostudies-literature | 2022 Jun

REPOSITORIES: biostudies-literature

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Publications

Linked-read sequencing for detecting short tandem repeat expansions.

Chiu Readman R Rajan-Babu Indhu-Shree IS Birol Inanc I Friedman Jan M JM

Scientific reports 20220607 1

Detection of short tandem repeat (STR) expansions with standard short-read sequencing is challenging due to the difficulty in mapping multicopy repeat sequences. In this study, we explored how the long-range sequence information of barcode linked-read sequencing (BLRS) can be leveraged to improve repeat-read detection. We also devised a novel algorithm using BLRS barcodes for distance estimation and evaluated its application for STR genotyping. Both approaches were designed for genotyping large ...[more]

PMID: 35672336

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Project description:Short tandem repeats (STRs), also known as microsatellites, are commonly defined as consisting of tandemly repeated nucleotide motifs of 2-6 base pairs in length. STRs appear throughout the human genome, and about 239,000 are documented in the Simple Repeats Track available from the UCSC (University of California, Santa Cruz) genome browser. STRs vary in size, producing highly polymorphic markers commonly used as genetic markers. A small fraction of STRs (about 30 loci) have been associated with human disease whereby one or both alleles exceed an STR-specific threshold in size, leading to disease. Detection of repeat expansions is currently performed with polymerase chain reaction-based assays or with Southern blots for large expansions. The tests are expensive and time-consuming and are not always conclusive, leading to lengthy diagnostic journeys for patients, potentially including missed diagnoses. The advent of whole exome and whole genome sequencing has identified the genetic cause of many genetic disorders; however, analysis pipelines are focused primarily on the detection of short nucleotide variations and short insertions and deletions (indels). Until recently, repeat expansions, with the exception of the smallest expansion (SCA6), were not detectable in next-generation short-read sequencing datasets and would have been ignored in most analyses. In the last two years, four analysis methods with accompanying software (ExpansionHunter, exSTRa, STRetch, and TREDPARSE) have been released. Although a comprehensive comparative analysis of the performance of these methods across all known repeat expansions is still lacking, it is clear that these methods are a valuable addition to any existing analysis pipeline. Here, we detail how to assess short-read data for evidence of expansions, reviewing all four methods and outlining their strengths and weaknesses. Implementation of these methods should lead to increased diagnostic yield of repeat expansion disorders for known STR loci and has the potential to detect novel repeat expansions.

Project description:BackgroundScreening for short tandem repeat (STR) expansions in next-generation sequencing data can enable diagnosis, optimal clinical management/treatment, and accurate genetic counseling of patients with repeat expansion disorders. We aimed to develop an efficient computational workflow for reliable detection of STR expansions in next-generation sequencing data and demonstrate its clinical utility.MethodsWe characterized the performance of eight STR analysis methods (lobSTR, HipSTR, RepeatSeq, ExpansionHunter, TREDPARSE, GangSTR, STRetch, and exSTRa) on next-generation sequencing datasets of samples with known disease-causing full-mutation STR expansions and genomes simulated to harbor repeat expansions at selected loci and optimized their sensitivity. We then used a machine learning decision tree classifier to identify an optimal combination of methods for full-mutation detection. In Burrows-Wheeler Aligner (BWA)-aligned genomes, the ensemble approach of using ExpansionHunter, STRetch, and exSTRa performed the best (precision = 82%, recall = 100%, F1-score = 90%). We applied this pipeline to screen 301 families of children with suspected genetic disorders.ResultsWe identified 10 individuals with full-mutations in the AR, ATXN1, ATXN8, DMPK, FXN, or HTT disease STR locus in the analyzed families. Additional candidates identified in our analysis include two probands with borderline ATXN2 expansions between the established repeat size range for reduced-penetrance and full-penetrance full-mutation and seven individuals with FMR1 CGG repeats in the intermediate/premutation repeat size range. In 67 probands with a prior negative clinical PCR test for the FMR1, FXN, or DMPK disease STR locus, or the spinocerebellar ataxia disease STR panel, our pipeline did not falsely identify aberrant expansion. We performed clinical PCR tests on seven (out of 10) full-mutation samples identified by our pipeline and confirmed the expansion status in all, showing absolute concordance between our bioinformatics and molecular findings.ConclusionsWe have successfully demonstrated the application of a well-optimized bioinformatics pipeline that promotes the utility of genome-wide sequencing as a first-tier screening test to detect expansions of known disease STRs. Interrogating clinical next-generation sequencing data for pathogenic STR expansions using our ensemble pipeline can improve diagnostic yield and enhance clinical outcomes for patients with repeat expansion disorders.

Dataset Information

Linked-read sequencing for detecting short tandem repeat expansions.

Publications

Linked-read sequencing for detecting short tandem repeat expansions.

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