Epitranscriptomic mapping of RNA modifications at single-nucleotide resolution using rhodamine sequencing (Rho-seq)

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Epitranscriptomics studies the layer of gene expression regulation through RNA modifications.
• Mapping RNA modifications is crucial for understanding gene expression but remains technically challenging.

Purpose of the Study:

• To introduce Rho-Seq, a novel protocol for mapping RNA modifications.
• To enable single-nucleotide resolution mapping of dihydrouridine and other RNA modifications.

Main Methods:

• Rho-Seq utilizes specific rhodamine-labeling of modified nucleotides.
• This labeling hinders reverse transcription, allowing for detection of modified sites.
• The protocol was optimized for dihydrouridine but tested for broader applicability.

Main Results:

• Rho-Seq successfully maps dihydrouridine RNA modification at single-nucleotide resolution.
• The method demonstrates applicability to other modifications, including 7-methylguanosine and 4-thiouridine.
• This expands the toolkit for epitranscriptomic analysis.

Conclusions:

• Rho-Seq provides a robust method for mapping RNA modifications.
• The technique offers high resolution and versatility for various modifications.
• This advancement facilitates deeper understanding of epitranscriptomics.