Identification and quantification of heterogeneously-methylated DNA fragments using epiallele-sensitive droplet digital polymerase chain reaction (EAST-ddPCR)

Publikation: Beitrag in FachzeitschriftForschungsartikelBeigetragenBegutachtung

Beitragende

Abstract

Background/Aim: DNA methylation plays an important role in the initiation and propagation of carcinogenesis; however, the role of heterogeneously methylated epialleles is currently not well studied, also due to the lack of sensitive, unbiased and high throughput methods. Here, a newly developed droplet digital PCR (ddPCR)-based method was evaluated regarding its ability to quantify such heterogeneously methylated epialleles with sufficient analytical sensitivity and specificity. Materials and Methods: Genomic DNA from blood leukocytes and bone marrow aspirate of an 8-year old male with B-cell acute lymphoblastic leukemia (B-ALL) and from normal and malignant prostate cell lines were analysed using ddPCR. Results: By using these DNA samples, the specificity of an applied set of fluorescence-labeled probes was demonstrated as a proof of concept. Conclusion: All individual heterogeneously-methylated epialleles were quantifiable by a set of fluorescence-labeled probes with complementary sequences to epialleles in a closed-tube and high-throughput manner. The new method named epiallele-sensitive droplet digital PCR (EAST-ddPCR) may give new insights in the generation and regulation of epialleles and may help in finding new biomarkers for the diagnosis of benign und malignant diseases.

Details

OriginalspracheEnglisch
Seiten (von - bis)299-312
Seitenumfang14
FachzeitschriftCancer Genomics and Proteomics
Jahrgang15
Ausgabenummer4
PublikationsstatusVeröffentlicht - 1 Juli 2018
Peer-Review-StatusJa

Externe IDs

PubMed 29976635
ORCID /0000-0002-3549-2477/work/148145340

Schlagworte

Ziele für nachhaltige Entwicklung

Schlagwörter

  • Digital PCR, DNA methylation, Heterogeneously methylated epialleles, Leukemia, Prostate cancer cells