Analysis of the RNA and Protein Complexome by Grad-seq

The complexome of a cell is the entirety of its complexes. Complexome capture studies have mostly focused on protein–protein interactions, which has left a gap in our knowledge of the global interactions of RNAs. To overcome these limitations, we recently introduced gradient profiling by sequencing (Grad-seq), which analyzes in a high-throughput fashion soluble cellular complexes after their separation in a glycerol gradient by fraction-wise RNA-seq and mass spectrometry. Here, we describe a detailed Grad-seq protocol for Streptococcus pneumoniae, which should also be applicable to other bacterial species.