Document Detail

Digital PCR analysis of maternal plasma for noninvasive detection of sickle cell anemia.
MedLine Citation:
PMID:  22451622     Owner:  NLM     Status:  MEDLINE    
BACKGROUND: Cell-free fetal DNA (cffDNA) constitutes approximately 10% of the cell-free DNA in maternal plasma and is a suitable source of fetal genetic material for noninvasive prenatal diagnosis (NIPD). The objective of this study was to determine the feasibility of using digital PCR for NIPD in pregnancies at risk of sickle cell anemia.
METHODS: Minor-groove binder (MGB) TaqMan probes were designed to discriminate between wild-type hemoglobin A and mutant (hemoglobin S) alleles encoded by the HBB (hemoglobin, beta) gene in cffDNA isolated from maternal plasma samples obtained from pregnancies at risk of sickle cell anemia. The fractional fetal DNA concentration was assessed in male-bearing pregnancies with a digital PCR assay for the Y chromosome-specific marker DYS14. In pregnancies with a female fetus, a panel of biallelic insertion/deletion polymorphism (indel) markers was developed for the quantification of the fetal DNA fraction. We used digital real-time PCR to analyze the dosage of the variant encoding hemoglobin S relative to that encoding wild-type hemoglobin A.
RESULTS: The sickle cell genotype was correctly determined in 82% (37 of 45) of male fetuses and 75% (15 of 20) of female fetuses. Mutation status was determined correctly in 100% of the cases (25 samples) with fractional fetal DNA concentrations >7%. The panel of indels was informative in 65% of the female-bearing pregnancies.
CONCLUSIONS: Digital PCR can be used to determine the genotype of fetuses at risk for sickle cell anemia. Optimization of the fractional fetal DNA concentration is essential. More-informative indel markers are needed for this assay's comprehensive use in cases of a female fetus.
Angela N Barrett; Thomas C R McDonnell; K C Allen Chan; Lyn S Chitty
Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2012-03-26
Journal Detail:
Title:  Clinical chemistry     Volume:  58     ISSN:  1530-8561     ISO Abbreviation:  Clin. Chem.     Publication Date:  2012 Jun 
Date Detail:
Created Date:  2012-05-29     Completed Date:  2012-07-31     Revised Date:  2014-03-14    
Medline Journal Info:
Nlm Unique ID:  9421549     Medline TA:  Clin Chem     Country:  United States    
Other Details:
Languages:  eng     Pagination:  1026-32     Citation Subset:  IM    
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MeSH Terms
Anemia, Sickle Cell / diagnosis*
DNA / blood,  genetics
DNA Mutational Analysis / methods*
INDEL Mutation
Polymerase Chain Reaction / methods
Polymorphism, Single Nucleotide
Prenatal Diagnosis / methods*
Sex Determination Analysis
Grant Support
RP-PG-0707-10107//Department of Health
Reg. No./Substance:

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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