Expression profiling by high throughput sequencing Methylation profiling by high throughput sequencing
Summary
Germ cells are vital for transmitting genetic information from one generation to the next and for maintaining the continuation of species. Here, we analyze the transcriptome of human primordial germ cells (PGCs) from the migrating stage to the gonadal stage at single-cell and single-base resolutions. Human PGCs show unique transcription patterns involving the simultaneous expression of both pluripotency genes and germline-specific genes, with a subset of them displaying developmental stage-specific features. Furthermore, we analyze the DNA methylome of human PGCs and find global demethylation of their genomes. Approximately 10-11 weeks after gestation, the PGCs are nearly devoid of any DNA methylation; with only 7.8% and 6.0% of the median methylation levels in male and female PGCs, respectively. Our work paves the way toward deciphering the complex epigenetic reprogramming of the germline with the aim of restoring totipotency in fertilized oocytes.
Overall design
In total, we analyzed the transcriptomes of 233 individual male and female human PGCs from 15 embryos at between 4 and 19 weeks of gestation as well as 86 neighboring somatic cells from 13 of these embryos using a single-cell RNA-Seq method that we developed. Furthermore, we analyzed the DNA methylomes of both male and female human gonadal PGCs as well as the neighboring somatic cells of eleven of these embryos at between 7 and 19 weeks of gestation using whole-genome bisulfite sequencing (WGBS). In total, we generated 1.3 billion 100 bp paired-end reads for transcriptome analyses and 4.1 billion 100 bp or 150 bp paired-end reads for DNA methylome analyses.
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