title: Improved single-cell protocol for preimplantation genetic diagnosis of spinal muscular atrophy.
abstract: OBJECTIVE
  To develop and validate a simple and reliable single-cell analysis protocol for the preimplantation genetic diagnosis (PGD) of spinal muscular atrophy (SMA).
  
  
  METHODS
  Molecular tests based on specific enzymatic digestion have already been described for SMA diagnosis. We modified the amplified DNA fragments so as to introduce a novel restriction site that provides an internal control for the completeness of the digestion.
  
  
  METHODS
  The genetics and reproduction departments of two teaching hospitals.
  
  
  METHODS
  Six informed couples at risk of transmitting SMA.
  
  
  METHODS
  All patients underwent standard procedures associated with intracytoplasmic sperm injection.
  
  
  METHODS
  Improvement of SMA diagnostic efficiency and accuracy on single cell.
  
  
  RESULTS
  One hundred fifty lymphocytes were analyzed with our protocol. One hundred percent diagnostic accuracy was achieved from both homozygous normal and SMN1-deleted leukocytes. Successful molecular analysis was achieved for 36 of 42 biopsied embryos (86%). Twenty-five normal embryos were transferred, but no pregnancy was achieved.
  
  
  CONCLUSIONS
  We developed an improved protocol for PGD of SMA that is simple, robust, and accurate; unfortunately, no pregnancies were achieved for any of the six patients who have undergone PGD in the program thus far.
authors.affiliation: Département de Génétique, U393, Hôpital Necker Enfants Malades, Paris, France.
authors.forename: Philippe
authors.initials: P
authors.lastname: Burlet
authors.forename: Nelly
authors.initials: N
authors.lastname: Frydman
authors.forename: Nadine
authors.initials: N
authors.lastname: Gigarel
authors.forename: Jean Paul
authors.initials: JP
authors.lastname: Bonnefont
authors.forename: Violaine
authors.initials: V
authors.lastname: Kerbrat
authors.forename: Gérard
authors.initials: G
authors.lastname: Tachdjian
authors.forename: René
authors.initials: R
authors.lastname: Frydman
authors.forename: Arnold
authors.initials: A
authors.lastname: Munnich
authors.forename: Julie
authors.initials: J
authors.lastname: Steffann
authors.forename: Pierre F
authors.initials: PF
authors.lastname: Ray
mesh_terms: D001483:Base Sequence; D004622:Embryo, Mammalian / Q000166:cytology / Q000502:physiology; D005260:Female; D005820:Genetic Testing / Q000379:methods / Q000706:statistics & numerical data; D006801:Humans; D008214:Lymphocytes / Q000166:cytology / Q000502:physiology; D008969:Molecular Sequence Data; D009134:Muscular Atrophy, Spinal / Q000175:diagnosis* / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000706:statistics & numerical data; D011247:Pregnancy; D019836:Preimplantation Diagnosis / Q000379:methods* / Q000706:statistics & numerical data