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Autosomal dominant polycystic kidney disease (ADPKD) is primarily characterized by renal cysts and progression to renal failure. It is a genetically heterogeneous disease, with mutations in the PKD1 gene accounting for the majority of cases. Direct mutation detection for PKD1-linked ADPKD or type 1 is complicated by the large size and complex genomic structure of PKD1. This paper describes a microsatellite marker-based assay for PGD in couples at risk of transmitting ADPKD type 1. During PGD, genetic analysis is carried out on single blastomeres biopsied from preimplantation embryos obtained after IVF, and only embryos unaffected by the disease under investigation are selected for transfer. Single-cell genetic analysis relied on a fluorescent duplex-PCR of linked polymorphic markers followed by fragment length determination on an automated sequencer. The co-amplification of the intragenic KG8 and the extragenic D16S291 marker at the single-cell level was evaluated in pre-clinical tests on lymphoblasts and research blastomeres. The developed assay proved to be efficient (96.1% amplification) and accurate (1.4% allele drop-out and 4.3% contamination), and can be applied in all informative ADPKD type 1 couples. From five clinical cycles carried out for three couples, two pregnancies ensued, resulting in the birth of two healthy children. © European Society of Human Reproduction and Embryology 2004; all rights reserved., SCOPUS: ar.j, info:eu-repo/semantics/published

Charcot-Marie-Tooth (CMT) disease is the 'common' name for a range of hereditary peripheral neuropathies. CMT1 is the most common form and is transmitted in an autosomal dominant manner. CMT1A maps to chromosome 17p11.2 and is caused, in the majority of cases, by a 1.5 Mb DNA duplication, that includes the peripheral myelin protein 22 (PMP) gene. This paper reports on preimplantation genetic diagnosis (PGD) for CMT1A in five couples. The CMT1A duplication was detected by fluorescent PCR analysis using polymorphic (CA)n markers localized within the duplication. Single-cell PCR on blastomeres allowed genetic analysis of embryos obtained after ICSI. Only healthy unaffected embryos were transferred to the uterus. PCR experiments with single EBV-transformed lymphoblasts or with research blastomeres allowed the evaluation of amplification efficiencies, as well as contamination and allele drop-out (ADO) rates for each PCR protocol. Three simplex PCR protocols (using one primer pair) and two duplex PCR protocols (using two primer pairs) were developed for CMT1A. Additionally, a protocol using all three primer pairs in triplex was also established. Thirteen clinical ICSI-PGD cycles were performed for five couples (12 simplex PCR cycles and one duplex PCR cycle), resulting in seven embryo transfers. Three singleton pregnancies ensued in two couples and three healthy babies were delivered. This report describes different fluorescent PCR-based tests which allow efficient and accurate single-cell level detection of the CMT1A duplication. On the basis of the presence of the healthy allele of the affected parent-to-be (and/or absence of the affected one), healthy embryos can be selected for transfer. The assays are suitable for PGD for other couples who present with the same CMT1A duplication [depending on their informativity for the (CA)n markers available] as described here., SCOPUS: ar.j, info:eu-repo/semantics/published

Cystic fibrosis (CF) is an autosomal recessive disease characterized by obstruction and chronic infections of the respiratory tract and pancreatic insufficiency. The gene was cloned in 1989 and the most frequent mutation was shown to be the ΔF508 mutation. During PGD, embryos obtained in vitro are checked for the presence or absence of the mutation, after which only embryos shown to be free of the mutation are returned to the mother. Up to 1999, 48 intracytoplasmic sperm injection (ICSI) and in vitro fertilization (IVF) cycles had been carried out for PGD for CF in 24 couples, and different diagnostic tests had been used to select non-affected embryos. Thirteen patients became pregnant and 12 healthy babies have been born. Copyright (C) 2000 John Wiley and Sons, Ltd., SCOPUS: ar.j, info:eu-repo/semantics/published

SCOPUS: ar.j, info:eu-repo/semantics/published