Nucleic Acid Capture from Human Blood Plasma Uncovers G-Quadruplex Structures
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…
The article details how researchers have discovered evidence of G-quadruplex structures within human blood plasma. Ultrashort cell-free DNA, which is about 50 nucleotides long, is present in human plasma but is difficult to detect using standard methods. Earlier studies suggested that this ultrashort DNA might be connected to regulatory regions and noncanonical structures, hinting at the potential role of higher-order DNA structures.
However, this idea had been based on computational predictions and not on solid experimental evidence.
In this study, scientists utilized computational, biophysical, and biochemical methods to test this hypothesis. They began by isolating ultrashort cfDNA from 20 healthy individuals and selectively enriching it at locations predicted to contain G-quadruplex sequences. These sequences were identified as both experimentally observed quadruplex regions and accessible chromatin in blood cells.
Using synthetic oligonucleotides corresponding to these enriched loci, the researchers observed that they tended to form parallel G-quadruplex structures, as confirmed through circular dichroism analysis. They also directly captured endogenous nucleic acids from pooled plasma using a method called poly(A)-tailing and immobilization, which avoided any extraction, denaturation, or annealing steps typically involved in processing DNA.
Two different probes, BG4 antibody and N-methyl mesoporphyrin IX, detected these G-quadruplex structures. To validate their findings, the team used a chemically unrelated G-quadruplex ligand called pyridostatin and found that it competed with the detected structures. This reciprocal competition provided strong confirmation of the signal.
In conclusion, this research provides strong experimental evidence that G-quadruplex structures are indeed present in human blood plasma, supporting the hypothesis that these higher-order DNA structures play a significant role in the unique properties of ultrashort cell-free DNA.
Written by urgent.news from bioRxiv's reporting — not their text. Machine-written — may contain errors; check the original before relying on it.
