Synthesis and characterization of N‐rich fluorescent bio‐dots as a reporter in the design of dual‐labeled FRET probe for TaqMan PCR: A feasibility study

DNA‐based analytical techniques have provided an advantageous sensing assay in the realm of biotechnology. Bio‐inspired fluorescent nanodots are a novel type of biological staining agents with excellent optical properties widely used for cellular imaging and diagnostics. In the present research, we...

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Bibliographic Details
Published in:Biotechnology and applied biochemistry 2023-04, Vol.70 (2), p.645-658
Main Authors: Imani, Mahsa, Mohajeri, Nasrin, Rastegar, Mojgan, Zarghami, Nosratollah
Format: Article
Language:English
Subjects:
Biocompatibility
bioconjugation
Biotechnology
bio‐dot
Carbodiimide
Coloring Agents
Conjugation
Design
DNA - genetics
DNA probes
Energy transfer
Feasibility Studies
Fluorescence
Fluorescence resonance energy transfer
Fluorescence Resonance Energy Transfer - methods
Fluorescent Dyes - chemistry
FRET probe
Oligonucleotides
Optical properties
Polymerase chain reaction
Probes
Quantum Dots
Sodium salts
Synthesis
Zeta potential
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Summary:DNA‐based analytical techniques have provided an advantageous sensing assay in the realm of biotechnology. Bio‐inspired fluorescent nanodots are a novel type of biological staining agents with excellent optical properties widely used for cellular imaging and diagnostics. In the present research, we successfully synthesized bio‐dots with excellent optical properties and high‐quantum yield from DNA sodium salt through the hydrothermal method. We conjugated the bio‐dots with 3' Eclipse Dark Quencher (Eclipse)‐labeled single‐strand oligodeoxyribonucleotide according to carbodiimide chemistry, to design a fluorescence resonance energy transfer (FRET) probe. The results confirmed the prosperous synthesis and surface functionalization of the bio‐dot. Analysis of size, zeta potential, and FTIR spectroscopy verified successful bioconjugation of the bio‐dots with probes. UV‐visibility analysis and fluorescence intensity profile of the bio‐dot and bio‐dot@probes represented a concentration‐dependent quenching of fluorescent signal of bio‐dot by Eclipse after probe conjugation. The results demonstrated that TaqMan PCR was not feasible using the designed bio‐dot@probes. Our results indicated that bio‐dot can be used as an efficient fluorescent tag in the design of fluorescently labeled oligonucleotides with high biocompatibility and optical features.
ISSN:0885-4513
1470-8744
DOI:10.1002/bab.2387