Work

Development of a His-Tag-mediated pull-down and quantification assay for G-quadruplex containing DNA sequences

Year: 2024

Type: article

Abstract: In this study, we developed a simple pull-down assay using peptide nucleic acids (PNAs) equipped with a His-Tag and a G-quadruplex (G4) ligand for the selective recognition and quantification of G4-forming DNA sequences. Efficient and specific target recovery was achieved using optimized buffer conditions and magnetic Ni-NTA beads, while quantification was realized by employing the enzyme-like properties of the G4/hemin complex. The assay was validated through HPLC analysis and adapted for a 96-well plate format. The results show that higher recovery can be achieved using His-Tag with Ni-NTA magnetic beads as compared to the more common biotin-streptavidin purification. The inclusion of the G4-ligand as an additional selectivity handle was shown to be beneficial for both recovery and selectivity.

Source: RSC Chemical Biology

DOI: 10.1039/d4cb00185k

Authors: Enrico Cadoni, Hanne Moerman, Annemieke Madder

Institution: Ghent University

Language: English

Topic: DNA and Nucleic Acid Chemistry

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: gold

Estimated APC paid: $1,962

Funders: Fonds Wetenschappelijk Onderzoek, Vlaamse regering