Prenatal genotyping of RHD and SRY using maternal blood

Area of Science:

• Molecular Biology
• Genetics
• Prenatal Diagnostics

Background:

• Non-invasive prenatal testing (NIPT) for fetal RHD genotyping is crucial for managing RhD-negative pregnancies.
• Polymerase chain reaction (PCR) techniques are widely used for fetal DNA analysis in maternal plasma.

Purpose of the Study:

• To evaluate a fluorescent duplex PCR method for fetal RHD genotyping in maternal plasma.
• To assess the impact of storage temperature on fetal DNA concentration in maternal blood samples.

Main Methods:

• Designed primers and probes for RHD exon 7 and the SRY gene.
• Performed monoplex and duplex PCR assays.
• Quantified fetal DNA in maternal plasma from 114 RhD-negative women.
• Investigated fetal DNA stability under different storage conditions.

Main Results:

• The duplex PCR showed reduced sensitivity for SRY gene amplification.
• An 8% discrepancy was observed between fetal RHD genotyping and postpartum serological typing.
• Fetal DNA concentration significantly decreased in samples stored at room temperature for 48 hours compared to those stored at <10°C.

Conclusions:

• The developed duplex PCR method is not suitable for routine clinical application due to sensitivity issues and lack of a positive control for RHD-negative female fetuses.
• Maintaining cool storage temperatures (<10°C) is essential for preserving fetal DNA integrity in maternal blood samples.