1. Generation of mouse–human chimeric embryos
- Author
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Boyang Zhang, Hanqin Li, Donald Yergeau, Aimee Stablewski, Houbo Jiang, Zhixing Hu, Brandon Marzullo, and Jian Feng
- Subjects
0303 health sciences ,Embryo ,Germ layer ,Biology ,Amplicon ,Embryonic stem cell ,General Biochemistry, Genetics and Molecular Biology ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,Mouse Embryonic Stem Cell ,Induced pluripotent stem cell ,030217 neurology & neurosurgery ,030304 developmental biology - Abstract
Naive human pluripotent stem cells (hPSCs) can be used to generate mature human cells of all three germ layers in mouse–human chimeric embryos. Here, we describe a protocol for generating mouse–human chimeric embryos by injecting naive hPSCs converted from the primed state. Primed hPSCs are treated with a mammalian target of rapamycin inhibitor (Torin1) for 3 h and dissociated to single cells, which are plated on mouse embryonic fibroblasts in 2iLI medium, a condition essentially the same for culturing mouse embryonic stem cells. After 3–4 d, bright, dome-shaped colonies with mouse embryonic stem cell morphology are passaged in 2iLI medium. Established naive hPSCs are injected into mouse blastocysts, which produce E17.5 mouse embryos containing 0.1–4.0% human cells as quantified by next-generation sequencing of 18S ribosomal DNA amplicons. The protocol is suitable for studying the development of hPSCs in mouse embryos and may facilitate the generation of human cells, tissues and organs in animals. The authors provide a protocol for generating mouse–human chimeric embryos by injecting naive human pluripotent stem cells converted from the primed state.
- Published
- 2021