Thor-Ribo-Seq: ribosome profiling tailored for low input with RNA-dependent RNA amplification

Translation regulation plays a pivotal role in the diversification of gene expression and the response to intra- and extracellular environmental cues. Ribosome profiling (or Ribo-Seq) serves as a sensitive, quantitative, comprehensive, and data-rich technique to survey ribosome traversal across the cellular transcriptome. However, due to the intricacy of library preparation, applications to low input have presented analytic challenges. To overcome this issue, here we developed Thor-Ribo-Seq, a ribosome profiling method tailored for low input. Thor-Ribo-Seq harnesses RNA-templated RNA transcription to linearly amplify ribosome footprints, assessing ribosome traversal at codon resolution with limited artifacts. This highly sensitized ribosome profiling approach provides a versatile option to investigate the translatome in precious samples.