A multiplex allele specific PCR capillary electrophoresis (mASPCR-CE) assay for simultaneously analysis of SMN1/SMN2/NAIP copy number and SMN1 loss-of-function variants

Objectives Spinal muscular atrophy (SMA) is a severe inherited neuromuscular disorder with a high carrier frequency and incidence rate. An accurate molecular method for SMA genes is crucial in carrier screening, clinical diagnosis, outcome assessment and precision therapies. Methods Comprehensively using the multiplex allele specific PCR (mASPCR) and capillary electrophoresis (CE), a novel single tube assay was developed to simultaneously determine the copy number of SMN1 / SMN2 / NAIP genes and five common loss-of-function variants in SMN1 . A total of 383 genotype known subjects were detected to evaluate the accuracy, while 564 clinical random samples were double-blind detected with this assay and MLPA to assess the specificity and sensitivity. Results This assay had high accuracy of 100 % consistency with the predetermined values in 383 genotype known subjects. Among 564 clinical random samples, the correlation between this assay and comparative method was 100 %, which showing high specificity and sensitivity. Conclusions This mASPCR-CE assay is easy to use and cost-effective, making it suitable for routine use in molecular screening and clinical diagnosis of SMA.