Detection of MicroRNA Expression Dynamics Using LNA/DNA Nanobiosensor.

Area of Science:

• Molecular Biology
• Cell Biology
• Biotechnology

Background:

• Investigating complex biological processes necessitates tools for probing spatiotemporal dynamics in individual cells.
• Current single-cell gene expression analysis methods like RNA in situ hybridization and single-cell PCR require cell lysis or fixation, preventing dynamic analysis.
• Real-time gene expression analysis in living cells remains a significant challenge.

Purpose of the Study:

• To develop a method for real-time single-cell gene expression analysis in living mammalian cells.
• To overcome the limitations of existing techniques that require cell lysis or fixation.
• To enable the study of dynamic and spatiotemporal gene expression regulation.

Main Methods:

• Development of a locked nucleic acid/DNA (LNA/DNA) nanobiosensor.
• The nanobiosensor utilizes a fluorophore-labeled detecting strand and a quenching strand.
• The probe hybridizes with target microRNA (miRNA), displacing the quencher and initiating fluorescence, detected via time-lapse microscopy.

Main Results:

• Demonstrated a method for single-cell gene expression analysis in living mammalian cells.
• Enabled large-scale dynamic monitoring of gene expression, including spatiotemporal distribution and heterogeneity.
• Showcased multiplex detection of miRNAs using different LNA/DNA nanobiosensors.

Conclusions:

• The LNA/DNA nanobiosensor provides a fast, generally applicable, and accessible method for real-time single-cell gene expression analysis.
• This technique facilitates the study of dynamic biological processes and gene expression heterogeneity in living cells.
• The method overcomes the limitations of traditional techniques by avoiding cell lysis or fixation.