松崎たちは6を怪しいと思ったから分析したんだよ。
>>
*The line subjected to WGS is indicated in parentheses in cases in which several sublines were established for one cell type. Other sublines were confirmed by PCR and sequencing.
Tsさんの最初の問題提起からね。
松崎たちはこの分析で具体的に何を分析したのかということね。
<STAP cells are derived from ES cells>のExtended Data Figure 2のCね。
Tsさんが言ってるのはAC129-1と129B6F1ES6とSTAP ChIP lysateの６番染色体の
SNIP比較が鮨されているが全部違ってるぞという指摘だわね。
Cの注は以下よ。内容的には桂報告にあることよね。
>>
c, Distribution of B6-type and 129-type SNPs along chromosome 6. The B6-homozygous SNP cluster (magenta) in the middle, which probably arose from the inheritance of the parental 129, is heterogeneous in length among six control ES cell lines. The four cag-GFP STAP stem-cell lines share the same length of the B6 SNP cluster with control ES 129B6F1 ES1. Note that the 129/B6-heterozygous SNP region in the 129 cag-GFP mouse is longer than that of AC129-1.
その支離滅裂な論理に対してTsさんはそれに構わず、図の結果だけに注目したのね。
①AC129-1
②129B6F1ES6
③STAP ChIP lysate
全部わずかに違ってるという指摘ね。そして①と③は同じだと言ってるんだろと
英語で書き込まれた。
>>
In Extended Data Figure 2.C, some differences are seen in Distribution of B6-type and 129-type SNPs between AC129-1 and STAP ChIP lysate.
Are these two cells the cells of the same individual?
そのSTAP ChIP lysate = 129B6F1ES1 の部分の説明は本文にあるのね。
>>
Control genomic DNA sequences for STAP cell chromatin immu-noprecipitation sequencing (ChIP-seq) experiments (Fig. 4 in ref. 2)had been deposited in the NCBI database2. To gain sufficient sequen-cing coverage, we re-sequenced the genomic DNA prepared from theSTAP cell lysate used for ChIP-seq (Extended Data Fig. 1a). We con-firmed that this STAP cell sample shared all the genomic character-istics described above for 129B6F1 ES1 (Extended Data Fig. 2c),indicating that the STAP cell sample used for ChIP-seq was derivedfrom 129B6F1 ES1 cells .
STAP細胞クロマチン免疫沈降シークエンシング（ChIP-seq）実験（Fig. 4 in ref. 2）のためのコントロールゲノムDNA配列はNCBIデータベースに登録されていた。 十分なシーケンシングカバレッジを得るために、我々はChIP-seqに使用したSTAP細胞溶液から調製したゲノムDNAを再配列決定しました（Extended Data Fig 1a）。 我々は、このSTAP細胞試料が、129B6F1ES1（Extended Data Fig. 2c）について上述した全てのゲノム特性を共有し、ChIP-seqに使用されるSTAP細胞試料が129B6F1 ES1細胞由来であることを示した。
そういうことなんだよ。そして更に、彼らは重大なごまかしをしている。
>>
しかも、肝心の129B6F1ES1 は、ゲノム解析されてないようで。
>The genome of 129B6F1 ES6 was sequenced, but further analysis showed that 129B6F1 ES1 rather than 129B6F1 ES6 shares all genomic anomalies found in AC129-1 (see main text).
本文からの引用文ではないのね。そこに対応する部分の本文は以下だわね。
>>
SNP analysis revealed that two independent STAP stem-cell lines,AC129-1 and AC129-2, had a 129B6F1 genetic background, whilethey were documented in the original article1as being establishedfrom 129 cag-GFP mice. We identified five heterozygous genomicanomalies: four deletions, and a duplication in these STAP stem-celllines (Extended Data Fig. 2b, d), which were not found in thesequenced parental mouse genomes. We identified that these anom-alies and sexual identity were shared by one of six control ES cell lineswith cag-gfp, 129B6F1 ES1, established earlier than AC129. This isalso the case for the other cag-GFP STAP stem-cell lines, FLS-T1 andT2, established in 2013. The 129B6F1 ES1 also shares a characteristichomozygous B6-SNP cluster in chromosome 6 with these four cag-GFP STAP stem-cell lines (Extended Data Fig. 2c, d). It is unlikely thatthe 129B6 ES1 line and these cag-GFP STAP stem-cell lines indepen-dently inherited all five chromosomal anomalies, the Y chromosome,and the same chromosome 6 from parental mice at establishment.
コピペで文字が繋がっちゃってるわね。やり直し。
>>
SNP analysis revealed that two independent STAP stem-cell lines,AC129-1 and AC129-2, had a 129B6F1 genetic background, while they were documented in the original article as being establishedfrom 129 cag-GFP mice. We identified five heterozygous genomicanomalies: four deletions, and a duplication in these STAP stem-celllines (Extended Data Fig. 2b, d), which were not found in the sequenced parental mouse genomes. We identified that these anomalies and sexual identity were shared by one of six control ES cell lines with cag-gfp, 129B6F1 ES1, established earlier than AC129. This is also the case for the other cag-GFP STAP stem-cell lines, FLS-T1 and T2, established in 2013. The 129B6F1 ES1 also shares a characteristic homozygous B6-SNP cluster in chromosome 6 with these four cag-GFP STAP stem-cell lines (Extended Data Fig. 2c, d). It is unlikely that the 129B6 ES1 line and these cag-GFP STAP stem-cell lines independently inherited all five chromosomal anomalies, the Y chromosome,and the same chromosome 6 from parental mice at establishment.
で、ここの論証自体は見事なんだけどね。肝心の129B6F1 ES1のデータが示されて
いないねとTsさんが指摘しているんだね。このSNIPと遺伝子異常はPCRで解析されていて
<肝心の129B6F1ES1 は、ゲノム解析されてないようで。>と言うのは事実で、WGSに
かけられたのは6だけだ。
>>
*The line subjected to WGS is indicated in parentheses in cases in which several sublines were established for one cell type. Other sublines were confirmed by PCR and sequencing.
とあって、表のカッコ内は6だ。
この中にヴァカンティが小保方さんの受け入れと自分の説の関係に言及しているところがあるわね。
>>
In 2002, Vacanti went to Brigham and Women’s as the head of anesthesiology, and opened a large tissue-engineering lab. Recognizing that the pursuit of stress-altered cells was well outside his area of expertise, he hoped to hire someone who could help. Several years later, a Japanese colleague mentioned that he had a talented student—at the top of her class in chemical engineering at Waseda University—who was looking for a tissue-engineering project. It was Haruko Obokata.
When Obokata arrived in Vacanti’s lab, he quickly recognized her as open-minded and astute. Thinking that she could add credibility and detail to his work on the sporelike cells, he asked her to recapitulate the study, employing the latest techniques in stem-cell research. For the time being, he withheld his hypothesis that harsh conditions could create stem cells. The last thing he wanted was for a graduate student from abroad to return home and develop the idea in someone else’s lab. His main concern, he told me, was: “Can we trust Haruko?”
⑥が昨日紹介したデイナ・グッドイヤーさんが記事に書いた部分ね。
<The last thing he wanted was for a graduate student from abroad
to return home and develop the idea in someone else’s lab.
His main concern, he told me, was: “Can we trust Haruko?”>
この部分はさすがにこの間の事情を調べた人でないと意味が分からないわよね。
確かに端折りすぎだわ。someone else’s labって若山研のことね。
そこは若山さんが提示した待遇条件はそれほどのものとは思えないんだよな。
手記は願ってもないことと書いてるけど、米国での待遇の方がいい筈なんだけどな。
ただ、自分の研究のためには今のGOFマウスの使える環境が欲しいんだね。そこのところは
手記は本音を書いてるんじゃないか。でも誘ってもらっているので若山研に行きたいと
行ったかどうかは怪しいよね。彼女は既にヴァカンティ研のポスドク契約をしているでしょ。
信義としてもおかしな話だ。それにヴァカンティがこの時考えていたのは
His main concern, he told me, was: “Can we trust Haruko?”ということ
だったんでしょ。つまり彼女を理研へ海外出張に出させて秘密保持は大丈夫かと心配した。
そうね。
>>
In the year since the papers’ retraction, he and Kojima had tweaked the procedure, making small adjustments to the media and returning to the original chemical stimulus, ATP, which also fuelled the cells, instead of the hydrochloric acid indicated in the publications. For months, they had tried to make a teratoma, the first major step to indicating pluripotency.
(続き)
>>
Now we walked over to Vacanti’s lab to see the results. He wasn’t optimistic. Before my arrival, he told me, he had called an emergency phone meeting with Kojima and his brother Martin. “I said, ‘We have to have something to demonstrate that this is indeed real or we’re going to look really stupid.’ ” As it happened, he said, just a few days before, he had seen the best-looking spheres in many months. They had tested positive for the embryonic-stem-cell markers Oct4 and Nanog, calculated by a machine that is not subject to the vagaries of auto-fluorescence. He had broken the spheres apart and placed the cells in a medium that encouraged the growth of neurons. The new plan was to test them for a protein called nestin, which, he said, would show that a mature skin cell had been transformed into a neural stem cell—taken back down to the root and up another branch.
(続き)
"Compact and taciturn, Kojima looked up and said “Good data” before saying hello. The nestin levels were fifty to sixty times as high as the last time he had checked. “Ha!” Vacanti said, jumping up and down and cheering.
“It’s only one sample,” Kojima said.
“Doesn’t matter,” Vacanti replied cheerfully. “One’s good for me.”
"
(続き)
In the progress of their research, they were all the way back to 2008, before Obokata, but Vacanti still saw a cause for celebration. After calling his brother, he decided that it was time for lunch and a glass of wine; Kojima came along, and Vacanti kept exhorting him to make merry. “I’m excited,” Kojima said. “But I need correct data, otherwise—” He chopped his hand through the air.
(続き)
Vacanti was reluctant to ruin a good day, but eventually he couldn’t put off looking at the teratoma slides any longer. Back in the lab, he sat down at the microscope and examined them for a long time. I stole a glance: a view of pink bubbles and marbled swirls like the endpapers of a seventeenth-century Venetian book. After a while, Vacanti concluded that the cells had failed to form new tissues; the only tissues evident had clearly come from the host mouse. Months of work, without producing evidence of pluripotency. But he didn’t dwell on that troubling thought for long. “Koji, you know what I was thinking?” he said. “We had Oct4, we had Nanog, we had nestin. Let’s never do another test. Let’s let it be someone else’s problem!”
ヴァカンティは最後に<Let’s let it be someone else’s problem!>と言ったと
デイナは書き留めていて、その実現が小保方さんのHPであるSTAP HOPE PAGEだわね。
そしてその上でヴァカンティは特許申請を推し進めることにした。それは利権でも
あると同時にその手続きを通して自分たちの捏造疑惑を払拭する手段でもあるのね。
勿論だよ。平たいところで実験したら増殖してシート状になる。それを
PGAに来るんでくちゃくちゃに丸めて皮下に埋めてるだけだから体細胞は
何らかの形で増殖しているさ。ヴァカンティが自分の目で確認したという
体細胞がそれだ。デイナは彼らがその時に使った細胞は皮膚の細胞と書いている。
でも、the only tissues evident had clearly come from the host mouse.だった。
これってドナー細胞の皮膚もあったんじゃないかな。ここのところはデイナ自身に
基礎知識が不足しているからはっきりしないね。
彼の<「STAP細胞事件」-STAP細胞もキメラもSTAP幹細胞もFI幹細胞も全てある>の
コメント欄のやり取りに関しては我々は東大はミトコンドリアの詳しい検証は
していないと思っている。
>>
Re: No title
FLSのミトコンドリアDNAがB6由来だったという話もありましたね。129B6だとミトコンドリアDNAは129由来になるはずで、だから核移植ではないかと疑われたと思います。これについては別の可能性もあると思っています。
2017.03.11 Sat l wamoga. URL l 編集
No title
・FLSは核移植ES細胞と予想して核移植ES細胞の全ゲノムを読んだが、どうやらFLSは核移植ES細胞ではなさそうだ。
・そこですでに得られたゲノムのデータでSTR解析をおこなった。
・受精卵ES細胞のFES1のみがFLSと一致した。
（日経サイエンス3月号の要約）
よくあることで珍しくはないんだけど、時が経過してもこんなにこじれてしまったのには別の
経緯があるのよね。
>>
Obokata resisted working on the stem-cell line; she wanted to remain focussed on the research she had been doing. When Wakayama pressed her, she grew embittered. In Boston, Kojima heard her screaming while reading her e-mail. He recalls that she shouted at the computer screen, “No! I don’t want to do it!” Kojima asked her what was going on. “Dr. Wakayama e-mailed me so many times,” she told him. “Like, ‘Did you do this assay? Did you do this experiment?’ Anything related to the stem-cell line, he forces me to do. I don’t want to, because I don’t know how to make it. I tried. I couldn’t.” In spite of their differences, Obokata writes, when Wakayama was offered a position at the University of Yamanashi he urged her to go with him.
和モガさんは丹羽さんが
>>
Klf4 expression was detected in all samples, which may reflect its expression in liver cells, and thus serves as a positive control in this assay.
