Characterization of the Epstein-Barr virus (EBV) DNA methylation status in blood circulating cell-free DNA.

Abstract

Background: Real time PCR detection of cell free circulating Epstein-Barr virus (EBV) DNA is a sensitive and specific tool to screen and to monitor the treatment of EBV associated malignancies. QIAamp® DNA Blood kit is widely used for DNA extraction despite its low extraction efficacy. This study aims to evaluate the efficacy of three commercial DNA extraction kits and to quantify methylation status of cell free circulating EBV DNA in nasopharyngeal carcinoma patients.

Methods: Extraction efficiency was evaluated by gel electrophoresis and real time PCR. Methylation status of cell free circulating DNA from twenty-one nasopharyngeal carcinoma patients was assessed by methylated DNA enrichment kit and real time PCR.

Results: Extraction efficiency of QIAamp® DNA Blood (QB), QIAamp® MinElute® Virus Spin (QV) and NucleoSpin® Plasma XS (NS) kits were 16.9% (±6.7%), 64.9% (±14.0%) and 0.53% (±0.43%) respectively. Both methylated and unmethylated EBV DNA were detected in NPC plasma.

Conclusion: QV was superior over QB and NS kits in efficiency of recovery of cell free circulating DNA. Quantifying DNA methylation levels is a potential biomarker to screen and monitor the treatment of EBV associated malignancies.