Hybrid molecular probe for nucleic acid analysis in biological samples

Area of Science:

• Biochemistry
• Molecular Biology
• Biomedical Research

Background:

• Intracellular nucleic acid monitoring is crucial for biomedical research.
• Molecular beacons (MBs) are common probes but suffer from false positives due to cellular processes.
• Existing MB limitations hinder accurate real-time, single-cell gene expression analysis.

Purpose of the Study:

• To design and synthesize a novel hybrid molecular probe (HMP) for reliable intracellular nucleic acid monitoring.
• To overcome the limitations of existing molecular probes, specifically false positives.
• To enable sensitive and selective detection of nucleic acid targets within living cells.

Main Methods:

• Designed and synthesized a hybrid molecular probe (HMP) composed of two DNA probes linked by a poly(ethylene glycol) (PEG) linker.
• One DNA probe is labeled with a donor fluorophore, and the other with an acceptor fluorophore.
• HMP utilizes proximity-induced fluorescence quenching and enhancement upon target binding.

Main Results:

• The HMP demonstrates high sensitivity, selectivity, and fast hybridization kinetics.
• HMP effectively prevents false positive signals caused by nuclease degradation, protein binding, or sticky-end pairing.
• The probe successfully detected nucleic acid targets from cellular samples with high specificity.

Conclusions:

• The developed HMP offers a robust solution for intracellular nucleic acid monitoring.
• HMP overcomes the false positive issues associated with traditional molecular beacons.
• This probe facilitates accurate real-time gene expression analysis at the single-cell level for biomedical applications.