A rapid and low-cost test for screening the most common Parkinson's disease-related GBA variants

Letizia Straniero, Valeria Rimoldi, Giada Melistaccio, Alessio Di Fonzo, Gianni Pezzoli, Stefano Duga, Rosanna Asselta

Research output: Contribution to journalArticlepeer-review


Introduction: Deleterious variants in the GBA gene confer a 2- to 20-fold increased risk of Parkinson's disease (PD) and are associated with a more severe disease course. The presence of a highly-similar pseudogene complicates genetic screening, both by Sanger and next-generation sequencing (NGS). Among PD-associated GBA variants, four missense substitutions (p.L444P, p.N370S, p.T369M, p.E326K) account for the majority of cases. Here, we aimed at developing an allele-specific PCR (AS-PCR) able to concomitantly detect the most common PD-related GBA variants. Methods: A multiplex PCR assay was designed using allele-specific oligonucleotides that distinguish the gene from the pseudogene and can exclusively amplify the variant alleles. Primer sequences and molarity, and thermal profiles were empirically optimized. The assay was validated on 4016 DNAs extracted by standard salting-out and previously analyzed by whole-exome sequencing (WES) followed by Sanger validation. Results: AS-PCRs performed on 4016 DNAs detected 103 variants; among them, 97 were true positives and 6 false positives. When comparing the results with the original WES data, for the “difficult” p.L444P, the number of false positives was 2/9 and 18/24 for multiplex-AS-PCR and WES, respectively. As we could have missed some p.L444P alleles by NGS, we verified the test performance on 50 DNAs from Sanger-validated p.L444P heterozygotes. All samples tested correctly. Conclusion: We set up and validated a rapid and inexpensive test for screening large cohorts of individuals for variants conferring a significant PD risk. This screening method is particularly interesting to identify patients who could benefit most from early access to personalized therapies.

Original languageEnglish
Pages (from-to)138-141
Number of pages4
JournalParkinsonism and Related Disorders
Publication statusPublished - Nov 2020


  • Allele-specific PCR
  • GBA
  • Genetic screening
  • Parkinson disease
  • Pseudogene

ASJC Scopus subject areas

  • Neurology
  • Geriatrics and Gerontology
  • Clinical Neurology


Dive into the research topics of 'A rapid and low-cost test for screening the most common Parkinson's disease-related GBA variants'. Together they form a unique fingerprint.

Cite this