Springer Protocols: Abstract: A Ribosomal Density-Mapping Procedure to Explore Ribosome Positions Along Translating mRNAs

A Ribosomal Density-Mapping Procedure to Explore Ribosome Positions Along Translating mRNAs

By: Naama Eldad³, Yoav Arava³

Abstract

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The number and distribution of ribosomes on a transcript provide useful information in ascertaining the efficiency of translation. Herein we describe a direct method to determine the association of ribosomes with specific regions of an mRNA. The method, termed Ribosome Density Mapping (RDM), includes cleavage of ribosomes-associated mRNAs with RNase H and complementary oligodeoxynucleotide followed by separation of the cleavage products on a sucrose gradient. The gradient is then fractionated and the sedimentation position of each mRNA fragment is determined by northern analysis. Although developed for yeast mRNAs, RDM is likely to be applicable to various other systems.

Affiliation(s): (3) Department of Biology, Technion - Israel Institute of Technology, Haifa, Israel

Book Title: Post-Transcriptional Gene Regulation

Series: Methods in Molecular Biology | Volume: 419 | Pub. Date: Dec-21-2007 | Page Range: 231-242 | DOI: 10.1007/978-1-59745-033-1_16

Subject: Genetics/Genomics

Key Words: Translation - mRNA - ribosome density - polysome - RNase H cleavage

References

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