Dataset Information

Expression data throughout reprogramming of MEF to iPS using a Dox-inducible promoter

ABSTRACT: 6C secondary MEFs were treated with Dox in mES media to turn on the Oct4, Klf4, cMyc, Sox2. Total RNA was extracted at day 0 (no Dox), day2, 5, 8, 11, 16 and 21 (with Dox) and day 30 (Dox-independent secondary iPS). RNA from Parental MEFs and Primary-iPS cells were also extracted for reference. 1B secondary MEFs were Dox treated for 5-days followed by RNA extraction. Subsequently, a culture removed of Dox treatment for an additional 5days was also analyzed.

ORGANISM(S): Mus musculus

PROVIDER: GSE21757 | GEO | 2010/07/01

SECONDARY ACCESSION(S): PRJNA127267

REPOSITORIES: GEO

ACCESS DATA

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Project description:The generation of induced pluripotent stem cells (iPSCs) by the forced expression of defined transcription factors in somatic cells holds great promise for the future of regenerative medicine. However, due to a poor understanding of the initial reprogramming mechanism, it is difficult to control the efficiency and quality of the iPSCs. Here we show that Zscan4, expressed transiently in 2-cell embryos and embryonic stem cells (ESCs), efficiently produces iPSCs from mouse embryo fibroblasts when coexpressed with Klf4, Oct4, and Sox2. Unlike other factors, Zscan4 is required only for the first few days of iPSC formation. Microarray analysis revealed transient and early induction of preimplantation-specific genes in a Zscan4-dependent manner. Our work indicates that Zscan4 is a previously unidentified potent natural factor that facilitates the reprogramming process through the recapitulation of the early embryonic program. Secondary MEFs were isolated from E13.5 embryos, which were harvested by tetraploid complementation. Secondary MEFs were plated on gelatin-coated 6-well plates at a density of 1x105 cells/well in the complete ES medium. After 24 h incubation, secondary MEFs were fed with complete ES medium with or without Dox (1.5 ?g/ml) and with or without 200 nM Tmx. Culture medium was changed every day. Withdrawal of drugs (Dox or Tmx) was always followed by 1x washing by PBS before changing culture medium. We cotransfected MEFs with PB-Tet-MKOS and either a PB-Tet-Zscan4 or PB-Tet-DsRed (control). These cells were subsequently cultured in the standard iPSC generation condition (Dox+). Unexpectedly, we found that MEFs transfected with a PB-Tet-Zscan4 show consistently higher efficiency of iPSC generation than MEFs transfected with a PB-Tet-DsRed (control). All selected iPSCs expressed pluripotent marker genes according to RT-PCR analysis and were positive for ALP, Nanog, and SSEA-1. These iPSCs also formed embryoid bodies in hanging drops without LIF and could be differentiated in vitro to cells of all three germ layers. We are currently studying whether the presence of Zscan4 has the beneficial effects on iPSCs in terms of genome stability.

Dataset Information

Expression data throughout reprogramming of MEF to iPS using a Dox-inducible promoter

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets