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Updated: May 6, 2026

An Easy Method for Plant Polysome Profiling

An Easy Method for Plant Polysome Profiling

Published on: August 28, 2016

Protocol for capturing the RNA-binding proteome from plants using orthogonal organic phase separation.

Victor A Sánchez-Camargo¹, Gertjan Kramer², Harrold A van den Burg³

¹Molecular Plant Pathology, Swammerdam Institute for Life Sciences, University of Amsterdam, 1098 XH Amsterdam, the Netherlands.

|November 29, 2025

Summary

This study introduces a new protocol to isolate RNA-binding proteins (RBPs) linked to RNA. This method aids in understanding RBP functions through proteomics analysis in plants.

Keywords:

Molecular biology Plant sciences Protein biochemistry

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Area of Science:

Molecular Biology
Proteomics
Plant Science

Background:

RNA-binding proteins (RBPs) are crucial regulators of RNA processing, including synthesis, function, transport, and decay.
Understanding RBP interactions with RNA is vital for deciphering gene regulation.
Existing methods for isolating RNA-protein complexes can be limiting.

Purpose of the Study:

To present a novel protocol for the mass isolation of RNA-binding proteins (RBPs) crosslinked to RNA.
To establish a strategy for subsequent proteomics analysis of these complexes.
To provide a method applicable to various plant tissues.

Main Methods:

In vivo UV crosslinking to stabilize RNA-protein interactions.
Tissue lysis and fractionation to prepare samples.
Purification of crosslinked RNA-protein adducts using organic solvents.
Proteomics analysis to identify bound proteins.

Main Results:

A comprehensive protocol for en masse isolation of crosslinked RNA-protein complexes was successfully developed.
The protocol enables subsequent proteomics analysis to identify RBPs.
Demonstrated applicability in Nicotiana benthamiana leaves, with potential for broader plant applications.

Conclusions:

The developed protocol offers an effective method for isolating RNA-binding proteins (RBPs) crosslinked to RNA.
This technique facilitates large-scale proteomics studies of RNA-protein interactions in plants.
The protocol is adaptable for diverse plant species and tissues, advancing RBP research.