Detection of SMN1 to SMN2 gene conversion events and partial SMN1 gene deletions using array digital PCR

Neurogenetics - Tập 22 - Trang 53-64 - 2021
Deborah L. Stabley1, Jennifer Holbrook1, Mena Scavina2, Thomas O. Crawford3,4, Kathryn J. Swoboda5, Katherine M. Robbins1, Matthew E. R. Butchbach6,7,8,9
1Nemours Biomolecular Core Laboratory, Nemours Biomedical Research, Nemours Alfred I. duPont Hospital for Children, Wilmington, USA
2Division of Neurology, Nemours Alfred I. duPont Hospital for Children, Wilmington, USA
3Department of Neurology, Johns Hopkins University, Baltimore, USA
4Department of Pediatrics, Johns Hopkins University, Baltimore, USA
5Department of Neurology, Center for Genomic Medicine, Massachusetts General Hospital, Boston, USA
6Center for Applied Clinical Genomics, Nemours Biomedical Research, Nemours Alfred I. duPont Hospital for Children, Wilmington, USA
7Department of Biological Sciences, University of Delaware, Newark, USA
8Department of Pediatrics, Thomas Jefferson University, Philadelphia, USA
9Center for Pediatric Research, Nemours Biomedical Research, Nemours Alfred I. duPont Hospital for Children, Wilmington, USA

Tóm tắt

Proximal spinal muscular atrophy (SMA), a leading genetic cause of infant death worldwide, is an early-onset motor neuron disease characterized by loss of α-motor neurons and associated muscle atrophy. SMA is caused by deletion or other disabling mutations of survival motor neuron 1 (SMN1) but retention of one or more copies of the paralog SMN2. Within the SMA population, there is substantial variation in SMN2 copy number (CN); in general, those individuals with SMA who have a high SMN2 CN have a milder disease. Because SMN2 functions as a disease modifier, its accurate CN determination may have clinical relevance. In this study, we describe the development of array digital PCR (dPCR) to quantify SMN1 and SMN2 CNs in DNA samples using probes that can distinguish the single nucleotide difference between SMN1 and SMN2 in exon 8. This set of dPCR assays can accurately and reliably measure the number of SMN1 and SMN2 copies in DNA samples. In a cohort of SMA patient–derived cell lines, the assay confirmed a strong inverse correlation between SMN2 CN and disease severity. We can detect SMN1–SMN2 gene conversion events in DNA samples by comparing CNs at exon 7 and exon 8. Partial deletions of SMN1 can also be detected with dPCR by comparing CNs at exon 7 or exon 8 with those at intron 1. Array dPCR is a practical technique to determine, accurately and reliably, SMN1 and SMN2 CNs from SMA samples as well as identify gene conversion events and partial deletions of SMN1.

Tài liệu tham khảo

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