Evaluation of extraction methods for methylated cell-free fetal DNA from maternal plasma.

Lim, Ji Hyae; Lee, Bom Yi; Kim, Jin Woo; Han, You Jung; Chung, Jin Hoon; Kim, Min Hyoung; Kwak, Dong Wook; Park, So Yeon; Choi, Hee Back; Ryu, Hyun Mee

Purpose: Recently, fetal placenta-specific epigenetic regions (FSERs) have been identified for quantification of cell-free fetal DNA (cff-DNA) for non-invasive prenatal testing (NIPT). The aim of this study was to evaluate the efficiencies of a column-based kit and magnetic bead-based kit for quantification of methylated FSERs from maternal plasma.Methods: Maternal plasma was extracted from normal pregnant women within the gestational age of 10~13 weeks (n = 24). Total cell-free DNA (cf-DNA) was extracted using a column-based kit and magnetic bead-based kit from the plasma of the same pregnant woman, respectively. Methylated FSERs were enriched from the extracted total cf-DNA using a methyl-CpG-binding domain-based protein method. The four FSERs were simultaneously quantified by multiplex real-time polymerase chain reaction.Results: Methylated FSERs were detected in all samples extracted from both kits. However, the amplification of FSERs showed significant differences in the extraction efficiency of methylated FSERs between the two extraction methods. The Ct values of methylated FSERs extracted using the column-based kit were significantly lower than those obtained using the magnetic bead-based kit (P P < 0.001 for all FSERs). Time and cost for the process of extraction were similar for the column kit and magnetic bead-based kit.Conclusions: Our findings demonstrate that the column-based kit was more effective than the magnetic bead-based kit for isolation of methylated FSERs from maternal plasma as assessed by FSER detection.

DNA methylation; ANEUPLOIDY; BIOMARKERS; PREGNANCY complications; PREVENTION of pregnancy complications; DIAGNOSIS

Journal of Assisted Reproduction & Genetics, 2018, Vol 35, Issue 4, p637

1058-0468

Academic Journal

10.1007/s10815-018-1114-8