Data Analysis Pipeline for scRNA-seq Experiments to Study Early Oogenesis

Germ cells as the means for the transmission of genetic information between generations have been a hot topic of research for decades. The analysis of the transcriptomes, that is of the RNA transcripts produced by the genotype at a given time, of germ cells and the surrounding somatic cells, is essential to unravel the cellular and molecular processes regulating gametogenesis. However, the asynchronized differentiation of germ cells and high cellular heterogeneity in the developing ovary or testis represent two unsurmountable challenges for delineating the transcription regulation mechanism of germ cells using traditional bulk RNA sequencing. By performing single-cell RNA sequencing (scRNA-seq), it is now possible to dissect the transcriptome of germ cell development at single-cell resolution, and apply powerful bioinformatics methods to translate raw sequencing data into meaningful information. Here, using the 10× Genomic platform and the most widely cited bioinformatics tools, we describe how to analyze early female germ cell development using scRNA-seq data generated from mouse E11.5 to E14.5 ovaries. This pipeline will provide a guide for exploring the processes of early germ cell development at single-cell resolution.