ccTaggedRP

Asynchronously cycling cells challenge the accurate characterization of phase-specific gene expression. Current strategies to characterize complex cell populations, including RNAseq, survey the steady state gene expression across the cell cycle and are inherently limited by the inability to resolve dynamic gene regulatory networks specific to subsets within the cell population. Single cell RNAseq (scRNAseq) provides an unbiased method to characterize the subpopulations and should identify different cell cycle transcriptomes if there are sufficient numbers of cycling cells present. Unfortunately, some cells are not amenable to scRNAseq. Therefore, we merged two powerful strategies, the incorporation of CDT1 and GMNN degrons used in FUCCI cell cycle sensors and the epitope-tagging of ribosomal proteins used in RiboTrap/Tag technologies, to create a cell-cycle dependent, tagged ribosomes, named ccTaggedRP. ccTaggedRPs are differentially expressed during the cell cycle, have similar subcellular locations as endogenous ribosomal proteins, and facilitate the recovery of cell cycle phase-specific RNA for sequencing. ccTaggedRP has broad applications to investigate phase-specific gene expression in complex cell populations. Examine cell cycle phase-specifiic RNA expression profiles via RNA-seqeuncing of mRNAs associated with ribosomomal proteins immunoprecipitated by either their HA epitope tag or Flag epitope tag