Specific detection of deleted and non‐deleted dystrophin exons together with gender assignment in preimplantation genetic diagnosis of Duchenne muscular dystrophy

• Published in: Molecular human reproduction 2003-07, Vol.9 (7), p.421-427
• Main Authors: Girardet, A., Hamamah, S., Déchaud, H., Anahory, T., Coubes, C., Hédon, B., Demaille, J., Claustres, M.
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Dystrophin; Exons; Gynecology. Andrology. Obstetrics; Humans; Key words: deletion/Duchenne muscular dystrophy/preimplantation genetic diagnosis/sex determination; Management. Prenatal diagnosis; Medical sciences; Muscular Dystrophy, Duchenne; Polymerase Chain Reaction; Pregnancy. Fetus. Placenta; Preimplantation Diagnosis
• ISSN: 1360-9947; 1460-2407; 1460-2407
• DOI: 10.1093/molehr/gag050

We have developed a preimplantation genetic diagnosis (PGD) strategy for Duchenne muscular dystrophy (DMD) allowing the simultaneous amplification of four exons (6, 8, 28 and 32) of the dystrophin gene together with ZFX/ZFY genes for gender determination. Preliminary experiments were carried out on 215 single lymphocytes from male and female individuals. Amplification rates ranged from 90.2% for exon 6 to 96.7% for exons 8 and 32. At least four of the five sequences were successfully amplified in 95.8% of single cells, and sexing was possible in 98.5%. This 5‐plex assay was found to be robust enough to be used in a PGD clinical procedure and was therefore applied to a family whose female partner was a heterozygous carrier of a large deletion extending from exon 21 to exon 34 of the dystrophin gene. We have thus analysed two exons located in the deleted region of the gene, two non‐deleted exons used as intrasample controls, and ZFX/ZFY genes. Cleavage stage embryo biopsy followed by PCR resulted in transfer of three unaffected embryos. The advantage of the present approach is to identify and subsequently transfer unaffected male embryos in addition to female embryos, and is now applicable to all families displaying a deletion involving at least one of these exons.