Non-invasive prenatal diagnosis of monogenic disorders: an optimized protocol using MEMO qPCR with miniSTR as internal control

Abstract : BACKGROUND: Analysis of circulating cell-free fetal DNA (cffDNA) in maternal plasma is very promising for early diagnosis of monogenic diseases. However, this approach is not yet available for routine use and remains technically challenging because of the low concentration of cffDNA, which is swamped by the overwhelming maternal DNA. METHODS: To make clinical applications more readily accessible, we propose a new approach based on mutant enrichment with 3'-modified oligonucleotides (MEMO) PCR along with real-time PCR to selectively amplify from the maternal blood the paternally inherited fetal allele that is not present in the maternal genome. RESULTS: The first proof of concept of this strategy was displayed for cystic fibrosis by the accuracy of our detection of the p.Gly542* mutation used as the initial developmental model. Subsequently, a retrospective study of plasmas originating from two pregnant women carrying a fetus with private mutation confirmed the effectiveness of our method. We confirmed the presence of cffDNA in the studied samples by the identification of a tri-allelic DNA profile using a miniSTR kit. CONCLUSIONS: This new non-invasive prenatal diagnosis test offers numerous advantages over current methods: it is simple, cost effective, time efficient and does not require complex equipment or bioinformatics settings. Moreover, our assays for different private mutations demonstrate the viability of this approach in clinical settings for monogenic disorders.
Type de document :
Article dans une revue
Liste complète des métadonnées

https://hal.umontpellier.fr/hal-02273004
Contributeur : Anthony Herrada <>
Soumis le : mercredi 28 août 2019 - 14:05:09
Dernière modification le : jeudi 29 août 2019 - 12:07:57

Identifiants

Collections

Citation

Claire Guissart, Vanessa Debant, Marie Desgeorges, Corinne Bareil, Caroline Raynal, et al.. Non-invasive prenatal diagnosis of monogenic disorders: an optimized protocol using MEMO qPCR with miniSTR as internal control. Clinical Chemistry and Laboratory Medicine, De Gruyter, 2015, 53 (2), pp.205-15. ⟨10.1515/cclm-2014-0501⟩. ⟨hal-02273004⟩

Partager

Métriques

Consultations de la notice

14