PMID: 11331673May 2, 2001Paper

Single cell multiplex PCR amplification of five dystrophin gene exons combined with gender determination

Molecular Human Reproduction
Pierre F RayArnold Munnich

Abstract

Large deletions in the dystrophin gene account for > 60% of mutations responsible for Duchenne muscular dystrophy (DMD). We have developed a genetic test that can be used directly for the preimplantation genetic diagnosis (PGD) of a majority of couples at risk of transmitting DMD. The test, a double nested multiplex polymerase chain reaction assay for the amplification of exons 8, 19, 45, 47 and 51 allows the detection of over 70% of all DMD deletions. Amelogenin sequences on the X and the Y chromosomes were also co-amplified to provide a correlation between embryo gender and deletion status. The setting up of reliable single cell assays for preimplantation genetic diagnosis is delicate and time consuming. Assays have to be validated on a large number of single cells for each specific mutation to assess efficiency and accuracy before being applied clinically. The multiplex procedure permitted the validation of all tested loci in the same series of isolated lymphocytes rather than in separate series for each exon. One hundred single lymphocytes, 50 female and 50 male cells, were analysed with an overall amplification rate of 98% and an amplification failure of 2% per exon. We suggest that this test is reliable, easy to set up an...Continue Reading

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Citations

Mar 7, 2007·Journal of Assisted Reproduction and Genetics·Akira NakabayashiYasunori Yoshimura
Feb 5, 2002·The Journal of Molecular Diagnostics : JMD·Alan R Thornhill, Karen Snow
Jan 5, 2006·European Journal of Human Genetics : EJHG·Nicola L DeanAsangla Ao
Jul 12, 2011·Stem Cell Reviews and Reports·Tatiana JazedjeMayana Zatz
Dec 25, 2009·Reproductive Biomedicine Online·A GirardetM Claustres
May 2, 2017·Expert Review of Molecular Diagnostics·Rebecca L Gould, Darren K Griffin

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