Development of an Effective Method to Enrich Cell-Free Nucleic Acids in Liquid Samples.

Abstract

Background: We aimed to establish a method to extract cell-free DNA (cfDNA) from liquid samples by combining protamine solution with nucleic acid extraction reagents.

Methods: Samples comprised 1 mL EDTA/2K plasma from healthy individuals (n = 22) and 5, 10, 20, and 40 mL K562 cell culture supernatants. After adding protamine solution and NaCl, the samples were incubated and subsequently centrifuged. Genomic DNA (gDNA) was extracted using a High Pure PCR Template Preparation Kit. For comparison, plasma was extracted using a cobas® cfDNA Sample Preparation Kit. The gDNA was subjected to real-time quantitative PCR.

Results: Our method yielded results comparable to those obtained using a cfDNA-specific kit. Culture supernatants showed volume-dependent yield variations, and gDNA was successfully extracted from samples of all volumes following enrichment.

Conclusions: Our method enhanced cfDNA enrichment, highlighting its potential for seamless integration with commonly available nucleic acid extraction reagents to process large sample volumes.