Targeted Profiling of RNA Translation

This unit describes a reverse transcription-quantitative PCR (RT-qPCR)-based method for gene-targeted measurement of RNA translation levels. In brief, the method comprises washing and lysis of cells with a buffer containing cycloheximide to enrich ribosomal accumulation at translation initiation sites (TIS), followed by enzymatic treatment to generate ribosomal footprints, reverse transcription targeted towards TIS of specific transcripts of interest to generate complementary DNA (cDNA), and qPCR to measure the abundance of these footprints. In this manner, this method enables time- and cost-effective assessment of changes in translation level across focused panels of genes and across numerous samples.