そこは本文に以下のように説明されているね。
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To confirm further that Fgf4-induced stem cells with a trophoblast-like nature were converted into ES-like cells, rather than just selecting ES-like cells pre-existing in the Fgf4-induced stem cell culture, we examined the effect of the MEK inhibitor PD0325901 on the ES-like cell generation from Fgf4-induced stem cells. Like trophoblast stem cells, Fgf4-induced stem-cell survival is dependent on FGF–MEK signals, and the inhibition of MEK activity caused massive cell death (Extended Data Fig. 6c). However, PD0325901 is also known to be a main effector in 2i medium[17] and to promote ES cell maintenance. Addition of PD0325901 to LIF+FBS-containing medium strongly inhibited the formation of ES-like colonies from Fgf4-induced stem cells (Fig. 3e, left, and Fig. 3f). This inhibition was unlikely to be due to secondary toxic effects from massive cell death of Fgf4-induced stem cells, as colonies formed in the presence of PD0325901 when ES cells were co-plated in the same culture with Fgf4-induced stem cells (Fig. 3e, right, and Fig. 3f).
単にFgf4誘導幹細胞の培養皿の中にあらかじめ存在していたES様細胞を選別してきたのではなく、栄養膜様の性質を有するFgf4誘導幹細胞がES様細胞に変換したのだということをさらに確認するために、我々は、Fgf4誘導幹細胞からES様細胞への生成に関して、MEK阻害剤、あるいはPD0325901の効果を調べた。栄養膜幹細胞のように、Fgf4誘導幹細胞の生存はFGF-MEKシグナルに依存しており、MEK活性を阻害すると大量の細胞死を引き起こした（拡張データ図6c）。しかし、またPD0325901は2i培地[17]の主要要素で、ES細胞の維持を促進することが知られている。 LIF + FBS含有培地にPD0325901を添加するとFgf4誘導幹細胞からのES様コロニーの形成を強く阻害した（図3e、左、および図3f）。 ES細胞がFgf4誘導幹細胞と同一の培養皿で共培養されていて、PD0325901の存在下でコロニーが形成されたのであるから、この阻害はFgf4誘導幹細胞の大量細胞死からの二次毒性効果の所為ではないようだ( 図3e、右、および図3f）。
FI-SCがLIF+FBS培地でES細胞様に変化したということの証明実験だよね。ここにそもそESが
コンタミしていたらいけないんで、そうでないということを証明しようとして、まず、FI-SCを
LIF+FBS培地に入れたものにMEK阻害剤を入れる。もしESがコンタミしていたらOct4-GFPが光る
はずだが光ってない。それが左側の図で、右側ではわざとOct4-GFPのES細胞を6%まぜている。
こちらは光っている。eのリジェンドは以下だ。
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e, No substantial formation of Oct4-GFP+ colonies was seen from Fgf4-induced cells in the presence of MEK inhibitor (left), whereas colonies frequently formed when cells were co-plated with Oct4-GFP ES cells (right; plated cells were 1/20 of Fgf4-induced cells).
e,MEK阻害剤（左）の存在下で、Fgf4誘導細胞からOct4-GFP陽性コロニーの実質的な形成は見られなかった。他方、同時播種したOct4-GFP ES細胞（右;播種した細胞はFgf4誘導細胞の1/20だった）からはコロニーが頻繁に形成された。
MEK阻害剤とは何かは本文にあるね。
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Like trophoblast stem cells, Fgf4-induced stem-cell survival is dependent on FGF–MEK signals, and the inhibition of MEK activity caused massive cell death (Extended Data Fig. 6c).
栄養膜幹細胞のように、Fgf4誘導幹細胞の生存はFGF-MEKシグナルに依存しており、MEK活性を阻害すると大量の細胞死を引き起こした（拡張データ図6c）。
(Extended Data Fig. 6c)の意味がよく分からないんだけどな。このリジェンドは
以下だ。
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c, MEK inhibitor treatment assay for Oct4-gfp Fgf4-induced stem cells in trophoblast stem-cell medium containing Fgf4. No substantial formation of Oct4-GFP+ colonies was observed from dissociated Fgf4-induced stem cells in MEK-inhibitor-containing medium. Scale bar, 100 μm.
c,Fgf4を含有する栄養膜幹細胞培地中のOct4-gfp陽性Fgf4誘導幹細胞に対するMEK阻害剤処理実験。 MEK阻害剤含有培地中の解離したFgf4誘導細胞から、Oct4-GFP陽性コロニーの実質的な形成は観察されなかった。 スケールバー、100μm。
荒食いの最後だ。冬に入ると食い渋る。今を逃して大釣りはないんだよ。
ああ、そうそう今序での余談だけど、
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33： 名無しさん ：2018/11/08(木) 10:08:43
『Failure to Replicate the STAP Cell Phenomenon』
「These results indicate that FI-SC samples are approximately 10% contaminated by TSC samples.」
丹羽検証論文のFig.3-aに下記条件でGFOの漏れ出しが報告された。
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a) Q-PCR analysis of the low-pH treated liver cells cultured for 7 days. Liver cells were prepared from 7-day old GOF mice and treated with either ATP or HCl, or
アーティクルに書かれているのは以下ね。
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The lack of substantial proliferation argues against the possibility that CD45− cells, contaminating as a very minor population in the FACS-sorted CD45+ cells, quickly grew and formed a substantial Oct4-GFP+ population over the first few days after the low-pH treatment. In addition, genomic rearrangements of Tcrb (T-cell receptor gene) were observed in Oct4-GFP+ cells derived from FACS-purified CD45+ cells and CD90+CD45+ T cells (Fig. 1i, lanes 4, 5, and Extended Data Fig. 2e–g), indicating at least some contribution from lineage-committed T cells. Thus, Oct4-GFP+ cells were generated de novo from low-pH-treated CD45+ haematopoietic cells by reprogramming, rather than by simple selection of stress-enduring cells.
マテメソにも以下のように書かれている。
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TCR-β chain gene rearrangement analysis
Genomic DNA was extracted from STAP cells and tail tips from chimaeric mice generated with STAP cells derived from CD45+ cells. PCR was performed with 50 ng DNA using the following primers (Dβ2: 5′-GCACCTGTGGGGAAGAAACT-3′ and Jβ2.6: 5′-TGAGAGCTGTCTCCTACTATCGATT-3′) that amplify the regions of the (D)J recombination. The PCR products were subjected to gel electrophoresis in Tris-acetate-EDTA buffer with 1.6% agarose and visualized by staining with ethidium bromide. PCR bands from STAP cells were subjected to sequencing analysis and identified as rearranged genomic fragments of the (D)J recombination.
ここだわ。
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Fig3
FI細胞をLIF+FBS培地で培養するとOct4、Nanog, SSEA-1を発現してきてES様細胞（STAP幹細胞）となり、このES様細胞は,TSマーカ（インテグリンα７、Eomes９）の発現を失う。
b ES様細胞はES並みの遺伝子発現（Oct4,Nalog,Rex1Kelf24,Sox2,Esrrb・・・）を保つが
TS並みの遺伝子発現（Cdx2、Eomes,、Elf5）を失う。
e f FI細胞はMEKインヒビターを入れて培養するとバラバラになって死んでしまうが、ES細胞は生き残る （TSとFIはFGF-MEKシグナル系に依存しているが、ESは依存していない）。
600： ペリー・メイスン ：2018/11/03(土) 17:45:51
リジェンドは以下だね。
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a, Culture of Oct4-GFP Fgf4-induced cells in LIF + 20% FBS medium.
b, qPCR analysis of ES-like cells derived from Fgf4-induced cells for pluripotent marker genes (left) and trophoblast marker genes (right). Values are shown as ratio to the expression level in ES cells (left) or trophoblast stem (TS) cells (right).
c, d, Culture of Oct4-GFP Fgf4-induced cells sorted by FACS for strong integrin α7 (Itga7) expression in LIF + 20% FBS medium.
d, Formation frequency (shown by percentage) of Oct4-GFP+ colonies from cells plated on gelatin-coated dishes at a clonal density. **P < 0.01; t-test; n = 3.
e, f, Culture of Oct4-GFP Fgf4-induced cells (dissociated) in LIF + 20% FBS medium with MEK inhibitor. **P < 0.01; NS, not significant; Tukey’s test; n = 3.
e, No substantial formation of Oct4-GFP+ colonies was seen from Fgf4-induced cells in the presence of MEK inhibitor (left), whereas colonies frequently formed when cells were co-plated with Oct4-GFP ES cells (right; plated cells were 1/20 of Fgf4-induced cells).
f, Quantification of colony formation per plated cells (1 × 103 Fgf4-induced cells and/or 1 × 103 ES cells). Unlike Fgf4-induced cells, ES cells formed colonies (regardless of co-plating with FI-SCs) in the presence of MEK inhibitor. Bars and error bars represent mean values and s.d., respectively (b, d, f). Scale bars: 100 μm (a, c, e).
え、それってバ感想の以下の質問に対する答えよね。
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32： 司書 ：2018/05/08(火) 19:10:31
(バ感想の英文)
Q. What are the genotypes of these ntES cells? Are these nuclear transferred ES cells from GOF (Oct4-GFP) mice?
