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Erschienen in:

2015 | Online First | Buchkapitel

Acquiring Ground State Pluripotency: Switching Mouse Embryonic Stem Cells from Serum/LIF Medium to 2i/LIF Medium

verfasst von : Matteo Tosolini, Alice Jouneau

Erschienen in: Methods in Molecular Biology™

Verlag: Springer New York

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Abstract

Mouse embryonic stem cells (ESCs) derive from the inner cell mass (ICM) of a blastocyst. These cells are pluripotent and thus able to generate both somatic and germinal lineages. It is possible to maintain ESCs in different pluripotent states depending on the in vitro culture conditions. Classically, ESCs are cultured in the presence of serum and LIF, which sustain the naive state of pluripotency but in this metastable state cells exhibit a large degree of heterogeneity. In the last few years, it has been discovered that when ESCs are cultured in a chemically defined medium (without serum), in the presence of LIF and with the addition of two small molecules (in particular the inhibitors of MAPK and Gsk-3 pathways), they reach a ground state of pluripotency where cells are more homogeneous and more “ICM-like.” In this protocol, we describe how we culture mouse ESCs and the way we switch them from naive to ground state.
Literatur
1.
Zurück zum Zitat Evans MJ, Kaufman MH (1981) Establishment in culture of pluripotential cells from mouse embryos. (Translated from eng). Nature 292(5819):154–156CrossRefPubMed Evans MJ, Kaufman MH (1981) Establishment in culture of pluripotential cells from mouse embryos. (Translated from eng). Nature 292(5819):154–156CrossRefPubMed
2.
Zurück zum Zitat Martin GR (1981) Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells. (Translated from eng). Proc Natl Acad Sci U S A 78(12):7634–7638PubMedCentralCrossRefPubMed Martin GR (1981) Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells. (Translated from eng). Proc Natl Acad Sci U S A 78(12):7634–7638PubMedCentralCrossRefPubMed
3.
Zurück zum Zitat Brook FA, Gardner RL (1997) The origin and efficient derivation of embryonic stem cells in the mouse. (Translated from eng). Proc Natl Acad Sci U S A 94(11):5709–5712PubMedCentralCrossRefPubMed Brook FA, Gardner RL (1997) The origin and efficient derivation of embryonic stem cells in the mouse. (Translated from eng). Proc Natl Acad Sci U S A 94(11):5709–5712PubMedCentralCrossRefPubMed
4.
Zurück zum Zitat Smith AG, Heath JK, Donaldson DD et al (1988) Inhibition of pluripotential embryonic stem cell differentiation by purified polypeptides. (Translated from eng). Nature 336(6200):688–690CrossRefPubMed Smith AG, Heath JK, Donaldson DD et al (1988) Inhibition of pluripotential embryonic stem cell differentiation by purified polypeptides. (Translated from eng). Nature 336(6200):688–690CrossRefPubMed
5.
Zurück zum Zitat Niwa H, Burdon T, Chambers I et al (1998) Self-renewal of pluripotent embryonic stem cells is mediated via activation of STAT3. (Translated from eng). Genes Dev 12(13):2048–2060PubMedCentralCrossRefPubMed Niwa H, Burdon T, Chambers I et al (1998) Self-renewal of pluripotent embryonic stem cells is mediated via activation of STAT3. (Translated from eng). Genes Dev 12(13):2048–2060PubMedCentralCrossRefPubMed
6.
7.
Zurück zum Zitat Brons IGM, Smithers L, Trotter M et al (2007) Derivation of pluripotent Epiblast Stem Cells from mouse and rat embryos. Nature 448:191–195CrossRefPubMed Brons IGM, Smithers L, Trotter M et al (2007) Derivation of pluripotent Epiblast Stem Cells from mouse and rat embryos. Nature 448:191–195CrossRefPubMed
8.
Zurück zum Zitat Tesar PJ, Chenoweth JG, Brook FA et al (2007) New cell lines from mouse epiblast share defining features with human embryonic stem cells. Nature 448:196–199CrossRefPubMed Tesar PJ, Chenoweth JG, Brook FA et al (2007) New cell lines from mouse epiblast share defining features with human embryonic stem cells. Nature 448:196–199CrossRefPubMed
9.
Zurück zum Zitat Wray J, Kalkan T, Smith AG (2010) The ground state of pluripotency. (Translated from eng). Biochem Soc Trans 38(4):1027–1032CrossRefPubMed Wray J, Kalkan T, Smith AG (2010) The ground state of pluripotency. (Translated from eng). Biochem Soc Trans 38(4):1027–1032CrossRefPubMed
10.
Zurück zum Zitat Marks H, Kalkan T, Menafra R et al (2012) The transcriptional and epigenomic foundations of ground state pluripotency. (Translated from eng). Cell 149(3):590–604PubMedCentralCrossRefPubMed Marks H, Kalkan T, Menafra R et al (2012) The transcriptional and epigenomic foundations of ground state pluripotency. (Translated from eng). Cell 149(3):590–604PubMedCentralCrossRefPubMed
11.
Zurück zum Zitat Habibi E, Brinkman AB, Arand J et al (2013) Whole-genome bisulfite sequencing of two distinct interconvertible DNA methylomes of mouse embryonic stem cells. (Translated from eng). Cell Stem Cell 13(3):360–369CrossRefPubMed Habibi E, Brinkman AB, Arand J et al (2013) Whole-genome bisulfite sequencing of two distinct interconvertible DNA methylomes of mouse embryonic stem cells. (Translated from eng). Cell Stem Cell 13(3):360–369CrossRefPubMed
Metadaten
Titel
Acquiring Ground State Pluripotency: Switching Mouse Embryonic Stem Cells from Serum/LIF Medium to 2i/LIF Medium
verfasst von
Matteo Tosolini
Alice Jouneau
Copyright-Jahr
2015
Verlag
Springer New York
DOI
https://doi.org/10.1007/7651_2015_207