Single-cell RNA sequencing and analysis of rodent blood stage Plasmodium.

Bulk RNA sequencing of Plasmodium spp., the causative parasite of malaria, fails to discriminate developmental-stage-specific gene regulation. Here, we provide a protocol that uses single-cell RNA sequencing of FACS-sorted Plasmodium-chabaudi-chabaudi-AS-infected red blood cells (iRBCs) to characterize developmental-stage-specific modulation of gene expression during malaria blood stage. We describe steps for infecting mice, monitoring disease progression, preparing iRBCs, and single-cell sequencing iRBCs. We then detail procedures for analyzing scRNA-seq data. For complete details on the use and execution of this protocol, please refer to Ramos et al.1.