Validation of Optical Genome Mapping for the Molecular Diagnosis of Facioscapulohumeral Muscular Dystrophy

The molecular diagnosis of facioscapulohumeral muscular dystrophy (FSHD) relies on detecting contractions of the unique D4Z4 repeat array at the chromosome 4q35 locus in the presence of a permissive 4q35A haplotype. Long, intact DNA molecules are required for accurate sizing of D4Z4 repeats. We vali...

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Bibliographic Details
Published in:	The Journal of molecular diagnostics : JMD Vol. 23; no. 11; pp. 1506 - 1514
Main Authors:	Stence, Aaron A., Thomason, Jon G., Pruessner, Jonathan A., Sompallae, Ramakrishna R., Snow, Anthony N., Ma, Deqin, Moore, Steven A., Bossler, Aaron D.
Format:	Journal Article
Language:	English
Published:	United States Elsevier Inc 01-11-2021 American Society for Investigative Pathology
Subjects:	Alleles Chromosome Aberrations Chromosome Mapping - methods Chromosomes, Human - genetics Cohort Studies Data Accuracy DNA - genetics DNA - isolation & purification Genetic Testing - methods Genome, Human Haplotypes Humans Molecular Diagnostic Techniques - methods Muscular Dystrophy, Facioscapulohumeral - blood Muscular Dystrophy, Facioscapulohumeral - diagnosis Muscular Dystrophy, Facioscapulohumeral - genetics Regular Reproducibility of Results Restriction Mapping - methods
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Summary:	The molecular diagnosis of facioscapulohumeral muscular dystrophy (FSHD) relies on detecting contractions of the unique D4Z4 repeat array at the chromosome 4q35 locus in the presence of a permissive 4q35A haplotype. Long, intact DNA molecules are required for accurate sizing of D4Z4 repeats. We validated the use of optical genome mapping to determine size and haplotype of D4Z4 alleles for FSHD analysis. The cohort included 36 unique DNA specimens from fresh blood samples or archived agarose plugs. High-molecular- weight DNA underwent sequence-specific labeling followed by separation and image analysis with data collection on the Saphyr system. D4Z4 allele sizes were calculated and haplotypes determined from the labeling patterns. Each specimen had previous diagnostic testing using restriction enzyme digests with EcoRI, EcoRI/BlnI, XapI, or HindIII, followed by pulsed field gel electrophoresis and Southern blot analysis with appropriate probes. Optical genome mapping detected 4q35 and 10q26 alleles ranging from 1 to 79 D4Z4 repeats and showed strong correlation with Southern blot allele sizing (R2 = 0.95) and haplotyping (133 of 134; 99.4% haplotype match). Analysis of inter-assay and intra-assay runs showed high reproducibility (0.03 to 0.94 %CV). Subsequent optical genome mapping for routine clinical testing from 315 clinical FSHD cases compared favorably with historical result trends. Optical genome mapping is an accurate and highly reproducible method for chromosomal abnormalities associated with FSHD.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 ObjectType-Undefined-3
ISSN:	1525-1578 1943-7811
DOI:	10.1016/j.jmoldx.2021.07.021