title: Single-tube nonaplex microsatellite PCR panel for preimplantation genetic diagnosis of Hb Bart's hydrops fetalis syndrome.
abstract: OBJECTIVE
  To develop a single-tube multi-marker assay for improved preimplantation genetic diagnosis (PGD) of deletional and/or non-deletional Hb Bart's hydrops fetalis syndrome, providing haplotype confirmation of deletional status, and maximization of linkage informativity.
  
  
  METHODS
  We performed in silico mining to identify novel microsatellites within 1 Mb flanking the alpha-globin gene cluster, and optimized a single-tube assay combining detection of α(0) -thalassemia deletions with multi-marker linkage analysis. We performed validation on 100 single cells prior to clinical PGD application.
  
  
  RESULTS
  Of 42 markers encompassing the α-globin gene cluster that were identified in silico, 9 were highly polymorphic (0.68 ≤ polymorphism information content ≤ 0.92; 0.66 ≤ Ho  ≤ 0.90; 10 ≤ alleles ≤ 35) and optimized to co-amplify directly from a single cell. A validation analysis of 100 single lymphoblasts yielded 100% amplification success for all markers, and individual marker allele drop-out (ADO) rates of 0-5%. Clinical application of the assay in PGD for Hb Bart's (2 cases/cycles) resulted in a twin pregnancy and healthy live birth of two baby girls.
  
  
  CONCLUSIONS
  This single-tube nonaplex microsatellite PCR panel can be applied directly to PGD of most deletional Hb Bart's without the need for deletion-specific customization, and to linkage-based PGD of non-deletional Hb Bart's.
authors: Yong Loo Lin School of Medicine, Department of Pediatrics, National University of Singapore, Singapore.
authors: Min
authors: M
authors: Chen
authors: KKIVF Center, Department of Reproductive Medicine, KK Women's and Children's Hospital, Singapore.
authors: Jerry K Y
authors: JK
authors: Chan
authors: KKIVF Center, Department of Reproductive Medicine, KK Women's and Children's Hospital, Singapore.
authors: Sadhana
authors: S
authors: Nadarajah
authors: Yong Loo Lin School of Medicine, Department of Pediatrics, National University of Singapore, Singapore.|Preimplantation Genetic Diagnosis Center, Khoo Teck Puat - National University Children's Medical Institute, National University Health System, Singapore.
authors: Arnold S C
authors: AS
authors: Tan
authors: KKIVF Center, Department of Reproductive Medicine, KK Women's and Children's Hospital, Singapore.
authors: Melinda L H
authors: ML
authors: Chan
authors: Clinic for Human Reproduction, Department of Obstetrics and Gynecology, National University Hospital, Singapore.
authors: Joyce
authors: J
authors: Mathew
authors: Preimplantation Genetic Diagnosis Center, Khoo Teck Puat - National University Children's Medical Institute, National University Health System, Singapore.
authors: Eugene E L
authors: EE
authors: Saw
authors: KKIVF Center, Department of Reproductive Medicine, KK Women's and Children's Hospital, Singapore.
authors: Cheryl
authors: C
authors: Lim
authors: KKIVF Center, Department of Reproductive Medicine, KK Women's and Children's Hospital, Singapore.
authors: Wendy
authors: W
authors: Wong
authors: Preimplantation Genetic Diagnosis Center, Khoo Teck Puat - National University Children's Medical Institute, National University Health System, Singapore.
authors: Felicia S H
authors: FS
authors: Cheah
authors: Department of Paediatric Medicine, KK Women's and Children's Hospital, Singapore.|Paediatrics Academic Clinical Program, Duke-NUS Graduate Medical School, Singapore.
authors: Hai-Yang
authors: HY
authors: Law
authors: Clinic for Human Reproduction, Department of Obstetrics and Gynecology, National University Hospital, Singapore.
authors: Peng-Cheang
authors: PC
authors: Wong
authors: Yong Loo Lin School of Medicine, Department of Pediatrics, National University of Singapore, Singapore.|Preimplantation Genetic Diagnosis Center, Khoo Teck Puat - National University Children's Medical Institute, National University Health System, Singapore.|Department of Laboratory Medicine, National University Hospital, Singapore.
authors: Samuel S
authors: SS
authors: Chong
mesh_terms: D000483:Alleles; D001483:Base Sequence; D002460:Cell Line; D003198:Computer Simulation; D004624:Embryo Transfer; D005260:Female; D005307:Fertilization in Vitro; D006239:Haplotypes; D006455:Hemoglobins, Abnormal / Q000235:genetics*; D006801:Humans; D015160:Hydrops Fetalis / Q000175:diagnosis / Q000235:genetics*; D007231:Infant, Newborn; D018895:Microsatellite Repeats; D008957:Models, Genetic; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D019836:Preimplantation Diagnosis / Q000379:methods*; D017384:Sequence Deletion; D017085:alpha-Thalassemia / Q000175:diagnosis / Q000235:genetics
chemical_list: D006455:Hemoglobins, Abnormal; C030448:hemoglobin Bart's