Quantifying telomeric lncRNAs using PNA-labelled RNA-Flow FISH (RNA-Flow).

Area of Science:

• Molecular Biology
• Genetics
• Biotechnology

Background:

• Long non-coding RNAs (lncRNAs) play crucial roles in cellular processes.
• Telomeric repeat-containing RNA (TERRA) and Telomerase RNA Component (TERC) are key lncRNAs associated with telomeres.
• Accurate quantification of lncRNAs is essential for understanding their function and for diagnostics.

Purpose of the Study:

• To present a novel, reliable method for detecting and quantifying specific lncRNAs.
• To enable the quantification of TERRA and TERC lncRNAs in various cell types.
• To establish a versatile tool for both basic research and clinical applications.

Main Methods:

• Development of the RNA-Flow method, combining peptide nucleic acid (PNA) probe specificity with flow cytometry.
• Application of the method to quantify cellular levels of TERRA and TERC lncRNAs.
• Adaptation of the staining protocol for confocal microscopy on unfixed cells.

Main Results:

• Successful detection and quantification of TERRA and TERC lncRNAs in cultured cell lines and peripheral blood mononuclear cells (PBMCs).
• Demonstration of the RNA-Flow method's ease of use and reliability.
• Validation of the protocol for lncRNA detection using confocal microscopy on unfixed cells.

Conclusions:

• The RNA-Flow method provides a robust and accessible approach for lncRNA quantification.
• This technique has broad applicability in molecular biology research and clinical diagnostics.
• The method facilitates the study of telomere-associated lncRNAs in various biological contexts.