Decreased DNA methylation of a CpG site in the HBAP1 gene in plasma DNA from pregnant women

OBJECTIVE: The objective of this study is to identify potential CpG site(s) or DNA methylation pattern(s) in the pseudo α-globin 1 gene (HBAP1 gene), the gene which locates in α-thalassemia-1 deletion mutation, to differentiate plasma DNA between pregnant and non-pregnant women. METHOD: DNA methylation profiles of placenta and peripheral blood from the MethBase database were compared to screen differentially methylated regions. This region was confirmed the differential by methylation-sensitive high resolution melt (MS-HRM) analysis. The differential region was used to compare DNA methylation profile of plasma DNA between pregnant and non-pregnant women by bisulfite amplicon sequencing in three levels: overall, individual CpG sites and individual molecules (DNA methylation patterns). RESULT: Using MethBase data, four consecutive CpG sites in the HBAP1 gene were identified as regions of differential DNA methylation between placenta and peripheral blood. The confirmation by MS-HRM showed the differential DNA methylation profile between the placenta and plasma from non-pregnant women. The comparison of DNA methylation profiles between the plasma of pregnant and non-pregnant women showed that, in the overall levels of the four CpG sites, DNA methylation of pregnant women was detected at lower levels than non-pregnant women. In the individual CpG site level, only the second CpG site showed differential DNA methylation between the groups. In the DNA methylation pattern level, there was no strongly significant differences in DNA methylation patterns between the pregnant and non-pregnant groups. CONCLUSION: Our result demonstrated that, in the plasma from pregnant women, only one of the four CpG sites displays a decrease in DNA methylation compared with non-pregnant women. It indicates that this CpG site might be useful for determining the presence or absence of fetal wild-type α-globin gene cluster allele in maternal plasma.