ArticleJournal of assisted reproduction and genetics2019
Evaluation and statistical optimization of a method for methylated cell-free fetal DNA extraction from maternal plasma.
Article in Journal of assisted reproduction and genetics, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 8 papers.
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Who cites it
8 citing papers in PubMed, 20 citations in OpenAlex.
- Preliminary Comparison of a Modified cfDNA Extraction Protocol for Y-Chromosome Marker Detection in Maternal Plasma.Diagnostics (Basel, Switzerland) · 2026Article
- sTLR9 on maternal Circulating immune cells as a negative regulatory phenotype during pregnancy.Cellular and molecular life sciences : CMLS · 2025Article
- Cartilage Repair in 2025: Hope, Hype, or Horizon?The archives of bone and joint surgery · 2025Article
- Ampholytic ion-exchange magnetic beads: a promising tool for selecting short fragments in circulating cell-free DNA analysis.Frontiers in oncology · 2024Article
- The impact of preanalytical variables on the analysis of cell-free DNA from blood and urine samples.Frontiers in cell and developmental biology · 2024Review
- Optimization of expression, purification and secretion of functional recombinant human growth hormone in Escherichia coli using modified staphylococcal protein a signal peptide.BMC biotechnology · 2021Article
- Impact of Preanalytical and Analytical Methods on Cell-Free DNA Diagnostics.Frontiers in cell and developmental biology · 2021Article
- Potential of Next-Generation Sequencing in Noninvasive Fetal Molecular Blood Group Genotyping.Transfusion medicine and hemotherapy : offizielles Organ der Deutschen Gesellschaft fur Transfusionsmedizin und Immunhamatologie · 2020Review
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Authors and funding
10 authors at 4 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
purposeMethylated cell-free fetal DNA (cffDNA) in maternal plasma can potentially be used as a biomarker for accurate noninvasive prenatal testing (NIPT) of fetal disorders. Recovery and purification of cffDNA are key steps for downstream applications. In this study, we aimed to developed and evaluated different aspects of an optimized method and compared its efficiency with common methods used for extraction of methylated cffDNA.
methodsSingle factor experiments, Plackett-Burman (PB) design, and response surface methodology (RSM) were conducted for conventional Triton/Heat/Phenol (cTHP) method optimization. The total cell-free DNA (cfDNA) was extracted from pooled maternal plasma using the optimized method called the Triton/Heat/Phenol/Glycogen (THPG), cTHP method, a column-based kit, and a magnetic bead-based kit. In the next step, methylated cfDNA from the extracted total cfDNA was enriched using a methylated DNA immunoprecipitation (MeDIP) kit. Real-time quantitative polymerase chain reaction was performed on the RASSF1 gene and hyper region to determine the genomic equivalents per milliliter (GEq/ml) values of the methylated cfDNA and cffDNA, respectively.
resultsThe optimum values of the significant factors affecting cfDNA extraction from 200 μl of plasma were 3% SDS, 1% Triton X-100, 0.9 μg/μl glycogen, and 0.3 M sodium acetate. The GEq/ml values of methylated cffDNA extracted using the THPG method were significantly higher than for the tested extraction methods (p < 0.001).
conclusionsOur results indicate that the THPG method is more efficient than the other tested methods for extraction of low copy number methylated cffDNA from a small volume of maternal plasma.
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