title: Preimplantation genetic diagnosis for familial amyloidotic polyneuropathy (FAP).
abstract: Preimplantation genetic diagnosis (PGD) was developed more than a decade ago to offer an alternative to prenatal diagnosis for couples at risk of transmitting an inherited disease to their offspring. Portuguese-type familial amyloidotic polyneuropathy (FAP type I), is an autosomal dominant disease presenting an inherited mutation in the gene encoding the plasma protein transthyretin (TTR). We here report the first protocol for single-cell detection of the Met30 mutation in FAP type I and its application to PGD. A nested PCR reaction for exon 2 of the TTR gene was developed. The PCR product was then analysed by restriction enzyme analysis and SSCP allowing the detection of the point mutation. Ten clinical cycles were performed in seven couples. From the 93 metaphase II (MII) injected oocytes, 82 were normally fertilized and 78 were biopsied. A positive signal in the nested PCR reaction was obtained in 61 blastomeres, corresponding to a DNA amplification efficiency of 78.2%. No allele dropout (ADO) or contamination were detected. A biochemical pregnancy was obtained in three cases and a clinical pregnancy in one couple is actually in normal evolution.
authors.affiliation: Department of Medical Genetics, Faculty of Medicine, University of Porto, Porto, Portugal. filipac@med.up.pt
authors.forename: F
authors.initials: F
authors.lastname: Carvalho
authors.forename: M
authors.initials: M
authors.lastname: Sousa
authors.forename: S
authors.initials: S
authors.lastname: Fernandes
authors.forename: J
authors.initials: J
authors.lastname: Silva
authors.forename: M J
authors.initials: MJ
authors.lastname: Saraiva
authors.forename: A
authors.initials: A
authors.lastname: Barros
mesh_terms: D028227:Amyloid Neuropathies, Familial / Q000175:diagnosis* / Q000235:genetics*; D001757:Blastomeres / Q000737:chemistry; D002478:Cells, Cultured; D004247:DNA / Q000032:analysis; D015252:Deoxyribonucleases, Type II Site-Specific; D004622:Embryo, Mammalian / Q000737:chemistry; D005260:Female; D006801:Humans; D008214:Lymphocytes; D008297:Male; D010062:Ovulation Induction; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D011174:Portugal; D011247:Pregnancy; D019836:Preimplantation Diagnosis; D020554:Sperm Injections, Intracytoplasmic; D020858:Tissue and Organ Harvesting
chemical_list: D004247:DNA; C048503:ATGCAT-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific