Switchable inhibitory behavior of divalent magnesium ion in DNA hybridization-based gene quantification

Hyowon Jin, Hyun Jeong Lim, Mark R. Liles, Beelee Chua, Ahjeong Son

Research output: Contribution to journalArticlepeer-review


Contrary to the understanding that divalent cations only result in under-estimation of gene quantification via DNA hybridization-based assays, we have discovered that Mg2+ could cause either under or over-estimation at different concentrations. Its switchable inhibitory behavior is likely due to its rigid first solvation (hydrated) shell and hence it is inclined to form non-direct binding with DNA. At low concentrations, it caused under-estimation by occupying the hybridization sites. At high concentrations, it caused probe, signaling and target DNA to aggregate non-specifically via Coulomb forces. By quantifying target DNAs at a range of Mg2+ concentrations using a gene quantification assay (NanoGene assay), a Mg2+ inflection concentration of ∼10−3 M was observed for both target ssDNA and dsDNA. Field emission scanning electron microscopy (FE-SEM), energy dispersive X-ray spectroscopy (EDS), and Fourier transform infrared spectroscopy (FT-IR) were employed to observe Mg2+-induced non-specific binding in the complexes that mimicked the presence of target DNA. Together with two other divalent cations Ca2+ and Cu2+, they were further examined via zeta potential measurements as well as NanoGene assay. This study revealed the importance of Mg2+ in achieving accurate gene quantification. Through a better mechanistic understanding of this phenomenon, it will be possible to develop strategies to mitigate the impact of Mg2+ on DNA hybridization-based gene quantification.

Original languageEnglish
Pages (from-to)4845-4856
Number of pages12
Issue number21
StatePublished - 18 Sep 2022

Bibliographical note

Funding Information:
This study was supported by the National Research Foundation of Korea (NRF-2019R1A2C2084233).

Publisher Copyright:
© 2022 The Royal Society of Chemistry.


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