Supplementary Figure 3: Early preimplantation stages progress in development during ESC derivation. | Nature Cell Biology

Supplementary Figure 3: Early preimplantation stages progress in development during ESC derivation.

From: The ability of inner-cell-mass cells to self-renew as embryonic stem cells is acquired following epiblast specification

Supplementary Figure 3

(A) Images of typical primary ESC colonies derived in 2i-LIF on gelatin from an E2.5 morula and the ICM of an E3.5 blastocyst after 6 and 5 days, respectively. (B) ESC derivation efficiencies from various developmental stages assayed by primary ES colony formation, as depicted in A. The numbers of embryos yielding an ESC colony per embryo are indicated. (C) Blastocoel formation of a morula 24 h after plating in 2i-LIF on gelatin. (D) PrE-like epithelial formation in an outgrowth from an E2.5 morula 4 days after plating in 2i-LIF on gelatin. We suspect that partial PrE specification is mediated by compensatory effects of LIF in 2i. (E) Surviving cells as judged by morphology at day 7 in percentage of wells per embryo after single ESC derivation in 2i on gelatin. The graph shows the mean and s.e.m. for the percentage of surviving cell-containing wells per embryo resulting from the analysis of n = ‘e’ embryos (with at least 3 wells analysed per embryo for each time point of embryonic development shown on the x axis, ‘w’ represents the total number of wells analysed per time point). (F) Images of surviving cells 7 days after single cell ESC derivation from E3.5 embryos in 2i-LIF on gelatin. (G,H) Single cell ESC derivation efficiency in serum-LIF (G) and under EpiSC conditions (ActA, bFGF) (H); Efficiency is displayed as the percentage of ESC-colony positive wells per embryo (ESC-col. pos. wells (%)/embryo) after 7 days. The total efficiency is further subdivided into ESC colonies arising from truly individual cells (ESC-col (single cell)) and ESC arising from small groups, usually between 2 and 5 cells (ESC-col (small group of cells)). The graphs show the mean and s.e.m. for the percentage of ESC colony-containing wells per embryo resulting from the analysis of n = ‘e’ embryos (with at least 3 wells analysed per embryo for each time point of embryonic development shown on the x axis, ‘w’ represents the total number of wells analysed per time point). Embryos were obtained from at least 3 individual litters on at least 2 different days.

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