Exploring translation regulation by global analysis of ribosomal association

Translation efficiency of an mRNA is related in most cases to its ribosomal association. This association can be readily measured through the separation of cellular complexes on sucrose gradients by velocity sedimentation, and identification of the sedimentation position of the mRNA in the gradient. Since ribosomes are the main driving force for mRNA sedimentation, sedimentation position is highly correlated with ribosomal association and thus translation efficiency. The advent of DNA microarrays allowed the determination of ribosomal association for many mRNAs in parallel through the combination of fractionation in a sucrose gradient followed by microarray analysis. This provided an enormous amount of novel information regarding translation control and regulation. Herein we provide a detailed protocol for performing such an analysis, indicating important points for consideration and discussing some of the advantages and limitations of this powerful approach.