Genome-wide analysis of mRNA polysomal profiles with spotted DNA microarrays

Abstract

The sedimentation of an mRNA in sucrose gradients is highly affected by its ribosomal association. Sedimentation analysis has therefore become routine for studying changes in ribosomal association of mRNAs of interest. DNA microarray technology has been combined with sedimentation analysis to characterize changes in ribosomal association for thousands of mRNAs in parallel. Such analyses revealed mRNAs that are translationally regulated and have provided new insights into the translation process. In this chapter, we describe possible experimental designs for analyzing genome-wide changes in ribosomal association, and discuss some of their advantages and disadvantages. We then provide a detailed protocol for analysis of polysomal fractions using spotted DNA microarrays.