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Updated: Jun 28, 2026

An Optimized Protocol for Electrophoretic Mobility Shift Assay Using Infrared Fluorescent Dye-labeled Oligonucleotides
Published on: November 29, 2016
Sequence dependent ultrafast electron transfer of nile blue in oligonucleotides
Rajib Kumar Mitra1, Sudarson Sekhar Sinha, Swati Maiti
1Unit for Nano-Science and Technology, Department of Chemical, Biological and Macromolecular Sciences, S.N. Bose National Centre for Basic Sciences, JD Block, Sector III, Salt Lake, Kolkata, 700098, India.
Abstract:
In this contribution we report studies on the nature of binding of nile blue (NB), a well known DNA intercalating drug, with three synthetic DNA oligonucleotides, (CGCAAATTTGCG)(2), (GCGCGCGCGCGC)(2) and (ATATATATATAT)(2). The nature of fluorescence quenching of the ligand upon complexation with the DNAs has been studied using steady state and picosecond-resolved optical spectroscopic techniques. The geometrical restriction on the probe in the DNA microenvironment is measured using picosecond-resolved rotational anisotropy measurements. Our experiments identify both non-specific electrostatic and intercalative modes of interaction of the probe with the DNAs at lower and higher DNA concentrations, respectively. This dual nature of binding is also confirmed through gel electrophoresis experiments. The nature of electron transfer (ET) reaction of GC base pairs with intercalated NB has also been explored. Competitive binding study reveals that binding affinity of the probe is higher with SDS micelles than with the DNAs within its structural integrity in presence of the micelles, as evidenced from circular dichroism (CD) measurements. The complex rigidity of NB with various DNAs and its fluorescence quenching with DNAs elucidate a strong recognition mechanism between NB and DNA.

