Nucleic Acid Capture from Human Blood Plasma Uncovers G-Quadruplex Structures
- 1Center for Molecular Medicine Cologne
Abstract
Ultrashort (US) cell-free DNA (cfDNA) is a population of approximately 50-nucleotide single-stranded DNA molecules in human plasma.1-3 Although it holds biological and diagnostic potential, US cfDNA escapes detection by conventional double-stranded library preparation methods.1-3 Independent studies have linked US cfDNA to regulatory genomic regions and to sequences predicted to form noncanonical structures, prompting the hypothesis that higher-order DNA structure contributes to its molecular properties. This interpretation, however, has so far rested on computational prediction rather than experimental evidence. Here we test this hypothesis using computational, biophysical and biochemical approaches. In silico size-selected US cfDNA from 20 healthy donors was selectively enriched at putative quadruplex sequences (PQS) that overlap both experimentally observed quadruplex sequences and accessible chromatin of blood cells. Synthetic oligonucleotides corresponding to the most enriched of these loci adopted predominantly parallel G-quadruplex (G4) structures, as revealed by circular dichroism. Endogenous nucleic acids captured directly from pooled plasma by poly(A)-tailing and immobilization, without extraction, denaturation or annealing at any step, displayed folded G4 structures. Two orthogonal probes detected these structures: the BG4 antibody and the fluorogenic ligand N-methyl mesoporphyrin IX. Reciprocal competition with a third, chemically unrelated G4 ligand, pyridostatin, confirmed the signal. Together, these experiments provide direct experimental evidence for the presence of folded G4 structures in human blood plasma.
Related
- Chapter 14: Unwinding Quadruplex Structures with RNA Helicases
- G-quadruplex structures affect the immune response to bacterial infection in mouse macrophages
- Promoter G-quadruplex structures regulate tissue-specific gene expression and functional differentiation in Bombyx mori
- Eps2Fold: a rapid method to characterize G-quadruplex DNA structures using single absorbance spectra
- Molecular origami in neuroscience: noncanonical DNA structures and functional nucleic acids in neural regulation and disease
BibTeX
@misc{gajarsky2026nucleic,
title = {{Nucleic Acid Capture from Human Blood Plasma Uncovers G-Quadruplex Structures}},
author = {M. Gajarsky and O. van Ray and T. Akkermann and P. Hunold and A. Cucchiarini and J.-L. Mergny and L. Trantirek and R. Haensel-Hertsch},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.05.747829},
url = {https://www.biorxiv.org/content/10.64898/2026.09.05.747829v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - Nucleic Acid Capture from Human Blood Plasma Uncovers G-Quadruplex Structures
AU - M. Gajarsky
AU - O. van Ray
AU - T. Akkermann
AU - P. Hunold
AU - A. Cucchiarini
AU - J.-L. Mergny
AU - L. Trantirek
AU - R. Haensel-Hertsch
PY - 2026
DA - 2026/09/05
PB - bioRxiv
DO - 10.64898/2026.09.05.747829
UR - https://www.biorxiv.org/content/10.64898/2026.09.05.747829v1
AB - Ultrashort (US) cell-free DNA (cfDNA) is a population of approximately 50-nucleotide single-stranded DNA molecules in human plasma.1-3 Although it holds biological and diagnostic potential, US cfDNA escapes detection by conventional double-stranded library preparation methods.1-3 Independent studies have linked US cfDNA to regulatory genomic regions and to sequences predicted to form noncanonical structures, prompting the hypothesis that higher-order DNA structure contributes to its molecular properties. This interpretation, however, has so far rested on computational prediction rather than experimental evidence. Here we test this hypothesis using computational, biophysical and biochemical approaches. In silico size-selected US cfDNA from 20 healthy donors was selectively enriched at putative quadruplex sequences (PQS) that overlap both experimentally observed quadruplex sequences and accessible chromatin of blood cells. Synthetic oligonucleotides corresponding to the most enriched of these loci adopted predominantly parallel G-quadruplex (G4) structures, as revealed by circular dichroism. Endogenous nucleic acids captured directly from pooled plasma by poly(A)-tailing and immobilization, without extraction, denaturation or annealing at any step, displayed folded G4 structures. Two orthogonal probes detected these structures: the BG4 antibody and the fluorogenic ligand N-methyl mesoporphyrin IX. Reciprocal competition with a third, chemically unrelated G4 ligand, pyridostatin, confirmed the signal. Together, these experiments provide direct experimental evidence for the presence of folded G4 structures in human blood plasma.
ER -