MicroRNA discovery and profiling in human embryonic stem cells by deep sequencing of small RNA libraries

We used massively parallel pyrosequencing to discover and characterize microRNAs (miRNAs) expressed in human embryonic stem cells (hESCs). Sequencing of small RNA cDNA libraries derived from undifferentiated hESC and from isogenic differentiating cultures yielded a total of 425,505 high-quality sequence reads. A custom data analysis pipeline delineated expression profiles for 191 previously annotated miRNAs, 36 novel miRNAs highly conserved across vertebrates, and 291 novel candidate miRNAs. A set of nine known and seven novel miRNAs were expressed in undifferentiated hESC and then strongly down-regulated with differentiation. Predicted mRNA targets of these miRNAs exhibited statistically significant enrichment for Gene Ontology functional categories relevant to ESC biology. Our study reveals that hESC express a larger complement of miRNAs than previously appreciated and it provides a resource for future studies of miRNA-mediated regulation in human embryonic stem cells. Two samples were 454 sequenced: undifferentiated human embryonic stem cells and differentiated human embryonic stem cells.