Prenatal diagnosis for CF using High Resolution Melting Analysis and simultaneous haplotype analysis through QF-PCR

Myrto Poulou; Aspasia Destouni; Georgia Kakourou; Emmanuel Kanavakis; Maria Tzetis

doi:10.1016/j.jcf.2014.04.002

Prenatal diagnosis for CF using High Resolution Melting Analysis and simultaneous haplotype analysis through QF-PCR

J Cyst Fibros. 2014 Dec;13(6):617-22. doi: 10.1016/j.jcf.2014.04.002. Epub 2014 Apr 29.

Authors

Myrto Poulou¹, Aspasia Destouni², Georgia Kakourou³, Emmanuel Kanavakis², Maria Tzetis³

Affiliations

¹ Department of Medical Genetics, Athens University, St Sophia's Children's Hospital, Athens 11527, Greece. Electronic address: mrpoulou@med.uoa.gr.
² 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.
³ Department of Medical Genetics, Athens University, St Sophia's Children's Hospital, Athens 11527, Greece.

PMID: 24784896
DOI: 10.1016/j.jcf.2014.04.002

Abstract

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.

Keywords: Cystic fibrosis; Haplotype analysis; High Resolution Melting; Mutation detection; Prenatal diagnosis; QF-PCR.

MeSH terms

Cystic Fibrosis / diagnosis*
Cystic Fibrosis / genetics
Female
Genetic Testing*
Genotyping Techniques / methods*
Haplotypes / genetics
Humans
Polymerase Chain Reaction / methods*
Pregnancy
Prenatal Diagnosis*
Reproducibility of Results
Retrospective Studies
Sensitivity and Specificity