Single-cell transcriptomics of small microbial eukaryotes: limitations and potential

Single-cell transcriptomics is an emerging research tool that has huge untapped potential in the study of microbial eukaryotes. Its application has been tested in microbial eukaryotes 50 mu m or larger, and it generated transcriptomes similar to those obtained from culture-based RNA-seq. However, microbial eukaryotes have a wide range of sizes and can be as small as 1 mu m. Single-cell RNA-seq was tested in two smaller protists (8 and 15 mu m). Transcript recovery rate was much lower and randomness in observed gene expression levels was much higher in single-cell transcriptomes than those derived from bulk cultures of cells. We found that the reason of such observation is that the smaller organisms had much lower mRNA copy numbers. We discuss the application of single-cell RNA-seq in studying smaller microbial eukaryotes in the context of these limitations.