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Updated: Feb 3, 2026
Assessment of Selective mRNA Translation in Mammalian Cells by Polysome Profiling
Published on: October 28, 2014
Polysome Profiling Analysis of mRNA and Associated Proteins Engaged in Translation
Eric S Pringle1,2, Craig McCormick1,2, Zhenyu Cheng1,2
1Department of Microbiology and Immunology, Dalhousie University, Halifax, Nova Scotia, Canada.
Abstract:
Post-transcriptional regulation is an important aspect of the control of gene expression. mRNAs are translated with variable efficiencies, and these efficiencies can change rapidly during adaptation to diverse environmental factors, including cellular stresses and microbial infections. Polysome profiling analysis utilizes ultracentrifugation to isolate complexes of mRNAs in the process of translation and corresponding proteins on the basis of density. Here we describe polysome profiling analysis using a continuous ultraviolet spectrophotometer and a gradient fractionator. We provide protocols for processing sucrose gradient fractions for isolation of RNA for RT-qPCR analysis and isolation of protein for SDS-PAGE and immunoblot analysis. © 2018 by John Wiley & Sons, Inc.
Insights
This study details polysome profiling, a method to analyze mRNA translation efficiency. It provides protocols for isolating RNA and proteins for further molecular analysis, aiding gene expression research.
Area of Science:
- Molecular Biology
- Gene Expression Regulation
Background:
- Post-transcriptional regulation is crucial for controlling gene expression.
- mRNA translation efficiency varies and adapts to environmental factors like stress and infection.
Purpose of the Study:
- To describe polysome profiling analysis for studying mRNA translation.
- To provide detailed protocols for subsequent molecular analyses.
Main Methods:
- Utilized ultracentrifugation to isolate translating mRNA-protein complexes based on density.
- Employed a continuous ultraviolet spectrophotometer and gradient fractionator.
- Developed protocols for RNA isolation (RT-qPCR) and protein isolation (SDS-PAGE, immunoblotting) from fractions.
Main Results:
- Established a comprehensive polysome profiling protocol.
- Demonstrated methods for analyzing RNA and protein components of translating mRNPs.
- Facilitated the study of dynamic changes in translation efficiency.
Conclusions:
- Polysome profiling is a valuable technique for investigating gene expression control at the post-transcriptional level.
- The described protocols enable detailed molecular characterization of translation complexes.
- This method supports research into cellular responses to environmental stimuli.
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01:31Translation
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