Determination of Fetal RHD Genotype Including the RHD Pseudogene in Maternal Plasma
- Resource Type
- Authors
- Eduardo Jens; Marcelo Zugaib; José Eduardo Levi; Alfredo Mendrone Junior; Carla Luana Dinardo; Adolfo Wenjaw Liao; Rossana Pulcineli Vieira Francisco; Marcia Regina Dezan; Karen Chinoca Ziza
- Source
- Subject
- Microbiology (medical)
Adult
Genotyping Techniques
Pseudogene
Clinical Biochemistry
Population
Prenatal diagnosis
030204 cardiovascular system & hematology
Biology
Umbilical cord
Sensitivity and Specificity
03 medical and health sciences
Exon
Young Adult
0302 clinical medicine
Pregnancy
Prenatal Diagnosis
Genotype
medicine
Immunology and Allergy
Humans
Prospective Studies
Allele
education
Research Articles
Fetus
education.field_of_study
030219 obstetrics & reproductive medicine
Rh-Hr Blood-Group System
Biochemistry (medical)
Public Health, Environmental and Occupational Health
Hematology
DNA
Molecular biology
Medical Laboratory Technology
medicine.anatomical_structure
Female
Pseudogenes
- Language
- English
Objective To examine the accuracy of fetal RHD genotype and RHD pseudogene determination in a multiethnical population. Methods Prospective study involving D-negative pregnant women. Cell-free DNA was extracted from 1 ml of maternal plasma by an automated system (MagNA Pure Compact, Roche) and real-time PCR was performed in triplicate targeting the RHD gene exons 5 and 7. Inconclusive samples underwent RHD pseudogene testing by real-time PCR analysis employing novel primers and probe. Results A positive result was observed in 128/185 (69.2%) samples and negative in 50 (27.0%). Umbilical cord blood phenotype confirmed all cases with a positive or negative PCR result. Seven (3.8%) cases were found inconclusive (exon 7 amplification only) and RHD pseudogene testing with both conventional and real-time PCR demonstrated a positive result in five of them, while two samples were also RHD pseudogene negative. Conclusion Real-time PCR targeting RHD exons 5 and 7 simultaneously in maternal plasma is an accurate method for the diagnosis of fetal D genotype in our population. The RHD pseudogene real-time PCR assay is feasible and is particularly useful in populations with a high prevalence of this allele.