Biogenesis of the mature bacterial ribosome requires the stepwise association of ∼50 ribosomal proteins and intricately folded rRNA, and involves ∼100 molecular chaperones. Each bacterium can generate ∼100,000 ribosomes per hour, and therefore the direct visualization of ribosome biogenesis has been challenging. Davis et al. genetically depleted the essential bL17 large subunit (LSU) ribosomal protein in Escherichia coli, and they then characterized the accumulated stalled assembly intermediates using chemical probing, mass spectrometry and single-particle cryo-electron microscopy. The authors identified 13 distinct LSU intermediates that were resolved to ∼4–5 Å, which revealed the arrangement of the intermediates in an assembly pathway in which blocks of structured rRNA and protein are cooperatively assembled. Assembly of mature ribosomes was shown to be flexible, with alternative assembly pathways being used when ribosomal components were limiting.