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. Author manuscript; available in PMC: 2019 Jul 1.
Published in final edited form as: Methods Enzymol. 2018 Aug 31;612:443–465. doi: 10.1016/bs.mie.2018.07.008

Figure 1.

Figure 1

Schematic of Caulobacter ribosome profiling procedure. Section 2.1 - Cell Growth, Synchronization and Harvesting: Grow 1 Liter of NA1000 cells, synchronize, and then grow to one of the cell cycle time points. Section 2.2 - Harvesting Cells: To each cell population, harvest by chloramphenicol pre-treatment (2 min at 28 °C) followed by rapid cell collection on ice. Section 2.3 - Cell Lysis: Lysis is performed under liquid nitrogen and a small portion is saved as a frozen aliquot for total RNA-seq. Section 2.4 - Footprinting and Footprint Extraction: Footprint mRNA fragments, select for size, and recover digested mRNA fragments. Section 2.5 - Total RNA-Seq Sample Preparation and PAGE Size Selection: Use the lysate aliquot from section 2.3 and prepare for total RNA-seq. Section 2.6 - Library Generation: Sequencing libraries are generated in the same way from the products of both ribosome footprinting and total RNA-Seq.