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Expression analysis in H9-hESCs to induce naive pluripotency


ABSTRACT: Human preimplantation embryo development involves complex dynamic cellular and molecular events that lead to the establishment of the three lineages of the blastocyst – the trophectoderm, primitive endoderm and epiblast. Owing to limited resources of biological specimens, our understanding of how the earliest lineage commitments are regulated is limited. Here, we examined role for MCRS1, TET1, and THAP11 in inducing naive pluripotency in human embryonic stem cells in vitro. Human embryonic stem cells (H9) were nucleofected with piggybac constructs that encode for three genes (MCRS1, THAP11, TET1) followed by neomycin selection for 2 weeks to ensure stable integration. Overexpression of genes can be induced by doxycycline (dox) administration to the culture medium. Prior to dox treatment cells were cultured in conventional feeder-free conditions and then transferred to a feeder layer. Culture medium was switched from mTeSR to W8. Administration of dox and media change into 2i/LIF (2 inhibitors against MEK and GSK3 pathway + leukemia inhibitory factor) induces the overexpression of piggybac constructs and over time transitions primed pluripotent hESCs into a naive pluripotent state. Both pluripotent states were examined with microarrays in three biological replicates for each condition. MCRS1 = microspherule protein 1; TET1 = tet methylcytosine dioxygenase 1; THAP11 = THAP domain containing 11

ORGANISM(S): Homo sapiens

PROVIDER: E-MTAB-4567 | BioStudies |

REPOSITORIES: biostudies

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