Prenatal diagnosis for CF using High Resolution Melting Analysis and simultaneous haplotype analysis through QF-PCR.
BACKGROUND
High Resolution Melting (HRM) Analysis is a validated, robust, low-cost, high throughput CF screening method. Here, we report the development and retrospective evaluation of the diagnostic value of a novel multiplex HRM, genotyping and haplotyping method for CF prenatal diagnosis (generic HRM/haplotyping).


METHODS
80 study samples from 20 carrier couples referred for PND (whole blood in EDTA and CVS or amniotic fluid) were genotyped retrospectively using the suggested protocol.


RESULTS
All DNA samples (variable sources, extraction methods and unknown concentrations) were successfully amplified by the 1st and 2nd round PCR. The Se, Sp, NPV and PPV for the generic HRM/haplotyping method are calculated at 100%.


CONCLUSIONS
This generic protocol for PND using HRM, facilitates the simultaneous analysis of DNA samples from various sources in a fast, robust and efficient way. It can be easily adapted and applied for any genetic condition.
Department of Medical Genetics, Athens University, St Sophia's Children's Hospital, Athens 11527, Greece. Electronic address: mrpoulou@med.uoa.gr.
Myrto
M
Poulou
Department of Medical Genetics, Athens University, St Sophia's Children's Hospital, Athens 11527, Greece; Research Institute for the Study of Genetic and Malignant Disorders in Childhood, St. Sophia's Children's Hospital, Athens 11527, Greece.
Aspasia
A
Destouni
Department of Medical Genetics, Athens University, St Sophia's Children's Hospital, Athens 11527, Greece.
Georgia
G
Kakourou
Department of Medical Genetics, Athens University, St Sophia's Children's Hospital, Athens 11527, Greece; Research Institute for the Study of Genetic and Malignant Disorders in Childhood, St. Sophia's Children's Hospital, Athens 11527, Greece.
Emmanuel
E
Kanavakis
Department of Medical Genetics, Athens University, St Sophia's Children's Hospital, Athens 11527, Greece.
Maria
M
Tzetis
D003550:Cystic Fibrosis / Q000175:diagnosis* / Q000235:genetics; D005260:Female; D005820:Genetic Testing; D060005:Genotyping Techniques / Q000379:methods*; D006239:Haplotypes / Q000235:genetics; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011247:Pregnancy; D011296:Prenatal Diagnosis; D015203:Reproducibility of Results; D012189:Retrospective Studies; D012680:Sensitivity and Specificity
Cystic fibrosis; Haplotype analysis; High Resolution Melting; Mutation detection; Prenatal diagnosis; QF-PCR