Project description:We report cell-type specific ribosome profiling in a mouse glioma model. We report a strategy for cell-type specific ribosome profiling in vivo. Our strategy was applied to the characterization of a mouse glioma model.
Project description:Ribosome profiling has emerged as a powerful tool for genome-wide measurements of translation, but library construction requires multiple ligation steps and remains cumbersome relative to more conventional deep sequencing experiments. We report a new approach to ribosome profiling that does not require ligation. Library construction for ligation-free ribosome profiling can be completed in one day with as little as 1 ng of purified RNA footprints. We used ligation-free ribosome profiling to identify new patterns of cell type-specific translation in the brain and tested its ability to identify translational targets of mTOR signaling in the brain.
Project description:We performed ribosome profiling (RIBO-seq) and transcriptome profiling (RNA-seq) to monitor RNAs associated with ribosome in the MCF-7 cell model Keywords: ribosome profiling, translation, MCF-7
Project description:Ribosome profiling has emerged as a powerful method to assess global gene translation, but methodological and analytical challenges often lead to inconsistencies across labs and model organisms. A critical issue in ribosome profiling is nuclease treatment of ribosome-mRNA complexes, as it is important to ensure both stability of ribosomal particles and complete conversion of polysomes to monosomes. We performed comparative ribosome profiling in yeast and mice with various ribonucleases including I, A, S7 and T1, characterized their cutting preferences, trinucleotide periodicity patterns, and coverage similarities across coding sequences, and showed that they yield comparable estimations of gene expression when ribosome integrity is not compromised. However, ribosome coverage patterns of individual transcripts had little in common between the ribonucleases. We further examined their potency at converting polysomes to monosomes across other commonly used model organisms, including bacteria, nematodes and fruit flies. In some cases, ribonuclease treatment completely degraded ribosome populations. Ribonuclease T1 was the only enzyme that preserved ribosomal integrity while thoroughly converting polysomes to monosomes in all examined species. This study provides a guide for ribonuclease selection in ribosome profiling experiments across most common model systems
Project description:We performed ribosome profiling of polysomes (Poly-RIBOseq) to monitor translation of RNAs associated with polysomes in brain extracts from a mouse model Keywords: ribosome profiling, translation, polysome profiling, Poly-RIBOseq
Project description:eIF2B5 is necessary for HrasG12V hyperplastic growth. Ribosome profiling was used to describe HrasG12V and WT dependence on eIF2B5-mediated translation.