Utilization of fluorescent nanomaterials as a tool to understand DNA secondary structures and implications toward DNA nanotechnology

Mukherjee, Ishani (2025) Utilization of fluorescent nanomaterials as a tool to understand DNA secondary structures and implications toward DNA nanotechnology. PhD thesis, Indian Institute of Science Education and Research Kolkata.

Text (PhD thesis of Ishani Mukherjee (17IP004)) 17IP004.pdf - Submitted Version Restricted to Repository staff only Download (9MB)

Abstract

The present thesis comprises synthesis and characterisation of fluorescent nanomaterials aimed to interact with DNA secondary structures, such as, i-motif, G-quadruplex, triplex DNA and tensegrity triangles to produce functional materials or composite systems having potential to cater to a broad range of DNA nanotechnology, bio-marking, therapeutics, etc. Neutral, small blue fluorescent carbon nanoparticles and a positively charged dye, (E)-3-ethyl-2-(4-(pyrrolidin-1-yl)styryl)benzo[d]thiazol-3-ium (EPSBT) that belongs to the hemicyanin family, formed a FRET pair. Carbon nanoparticles, with the help of dipole orientation and distance dependent FRET decoded the binding location of EPSBT in different single stranded DNA sequences. Carbon nanoparticles derived from single precursor, for example, folic acid, interacted with biologically important C-rich i-motif DNA sequences. It resulted in destabilisation of the i-motif structures with varying slightly for intra and intermolecular i-motifs. This showed the potential therapeutic plausibility of the folic acid derived carbon nanoparticles. C-rich and G-rich sequences are complementary of each other. Using the partial complementarity of a G-rich and a C-rich sequence, a stimuli responsive DNA network was synthesized. The G-rich G4 forming sequence was tagged with a blue fluorescent carbon nanoparticle and a C-rich i-motif forming sequence with gold nanoparticles. Mixing these two DNA coated nanoparticles in proportions, generated μM sized supramolecular assemblies. Carbon and gold nanoparticles formed a donor-quencher pair which helped in investigating the assembly formation spectroscopically. Since C-rich forming sequences are proton aptamers, the assembly showed a network tearing at pH 5 and a 3D network compaction upon increasing the pH to 8. Next, a 2T7 tensegrity template was assembled from ss-DNAs and silver nanoclusters were synthesised in the developed pockets. The excited state lifetime of the tensegrity triangle templated clusters increased by ~2 ns from 6.5 ns to 8.3 ns upon adding short oligonucleotides complementary to the 2T7 inter-junction and arm sequences. This is because the short oligonucleotide provides further rigidity to the 2T7 template by forming triads with its ds-DNA. This also alleviates the issues of forming nanoclusters using ds-DNA.

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