私たちは、胚の分離の彼らのアドバイスをタルNawyとマーティンバイエルに感謝したいと思います。 MTR、VG、UGおよびCBは、胚分離装置を考案した。 MTR、VGとCBが胚分離プロトコルを開発しました。 MTR、VGとCBは、プロトコルを確立し胚を単離し、胚cDNAを生成し、VGは、PCR、MTR GUS染色、JJ FISH実験を行った。 MTR、VG、CGとUGは、原稿を書いた。この作品は、チューリッヒ大学、ロシュ研究財団（MTR）に旅の仲間、そしてスイスの建国（UGとCBへ）からの補助金によって賄われていた。

手順は、 図1に示すフローチャートに要約されている。マイクロキャピラリーや楽器のセットアップを図2及び図3に示されており、胚の分離の一般的な手順を図4に示す。 1。材料およびバッファの準備ガラスマイクロキャピラリーの1.1シリコンコーティング<ol><li> Sigmacote（シグマ）の〜5mlで15ミリリットルファルコンチューブにマイクロキャピラリーを置き、数回転倒。 </li><li>溶液を除去し、60℃でアルミホイルで毛細血管を含むファルコンチューブを配置し、3時間のためにそれらを焼く室温で保管してください。 </li></ol> 1.2〜50〜100μmの直径のマイクロキャピラリーチップを入手<ol><li>手動でブンゼンバーナーの上または商業プラー（垂直フィラメントプラーまたはマイクロピペットプラー）を使用して、どちらか1ミリメートル口径ガラスキャピラリーを引き出します。 </li><li>ダイヤモンドを使用してください先端のペンまたは所望の開口部を作成するために引っ張らキャピラリーの先端を切断する刃。実体顕微鏡下で最高の形の毛細血管を選択します。開口部は、50〜100ミクロン（ 図2）である必要があります。 </li></ol> 1.3スライドの準備<ol><li> 5分間Sigmacote（〜1ミリリットル/スライド）を持つすべての井戸を覆うことにより、クリーンなスライドをシリコーンで覆う、取り外します。アルミホイルで焼く3時間。室温で保管してください。 </li><li> 10％SDS、ヌクレアーゼフリー水（DEPC-のddH 2 Oをオートクレーブ）、70％エタノールで2分、100％エタノールで2分間、空気中で2×2分で10分間のマルチウェルガラス顕微鏡スライドを洗う乾いた。すべてのステップはオートクレーブコプリンジャーで行われます。スライドは、各使用の間の徹底したクリーニングを提供することを複数回再利用することができる。 </li><li>直前胚分離普及〜100.5μlのmg / mlのウシ血清アルブミン（BSA）を各ウェルと空気乾燥の全面ピペットで。 </li></ol> 1.4顕微鏡とキャピラリセットアップ &lt;OL&gt; <li> 10倍と20倍の倍率を目的とした倒立顕微鏡を使用してください。光のコントラストを最適化する（胚はかなり透明である）。 </li><li>顕微鏡の横にあるガラスキャピラリーを保持するために、マイクロマニピュレータを配置します。キャピラリーガラスがゴムチューブを介してマイクロインジェクター（ 図3A）または正規のP-200ピペットに接続されている（ 図3B、このセットアップの詳細については説明を参照）。 </li><li> 〜70°の角度で顕微鏡スライド（ 図3）の上にキャピラリーを配置し、視野内の開口部を持つ位置を調整します。 </li><li>直前の胚の分離にコート毛細血管までかかると放し〜5-10μlのBSA（1 mg / ml）で。 </li></ol> 1.5バッファ 表1は、下流のアプリケーションに応じて分離し、先のバッファを示しています。 <ol><li> 〜サンプル当たり1ミリリットル分離バッファが新たに前に使用し、氷の上でそれを維持する準備。 </li><li>氷（分子アプリケーション）または湿潤チャンバー内顕微鏡スライド（細胞学的アプリケーション）で0.5ミリリットルエッペンドルフ低結合チューブに宛先バッファを準備します。 </li></ol> 2。シード解剖と胚抽出種子開発の2.1同期<ol><li class="jove_content">花を去勢し、他の花で露光雌しべとの接触を回避しながら、成長室に2日間保管してください、その後、（ 例えば 11）、それらを受粉。 </li><li class="jove_content">クリア種子の微視的調査することにより、成長条件の下で開発の段階をテストします。当社の成長条件で（21で16時間明°C、18暗所8時間℃、70％湿度）種子は2.5日受粉後に収集（DAP）は、主に2-4細胞胚および種子は3.5収集もたらした - 4 DAPは球状もたらし胚。 </li></ol> 2.2シード解剖と破裂<ol><li> Sを削除鉗子とインスリン針で実体顕微鏡下10-15長角果（〜2.5 DAP）からeeds。 </li><li>氷上に置き2ミリリットル丸底エッペンドルフチュー​​ブに20μlの分離バッファ内の種子を浸す。 </li><li>穏やか種子抽出物が濁っなるまで胚を放出するプラスチック乳棒（DEPC-のddH 2 Oで洗浄し、70％エタノールで洗浄し、10％SDSで洗浄済み）を用いて種子をつぶす。適用するための力は、トライアルの際、すべてのユーザによって決定されるべきである。 </li><li>杵を洗って、サンプルを希釈する分離バッファ300μlのと乳棒をすすぐ。 </li><li>スピンダウン5秒5,000×gで抽出。優しく上下2〜3回ピペッティングしてペレット化エキスを再懸濁。 </li><li> 30μmのナイロンメッシュ（PartecCelltricks などから、チューブアダプタに取り付けられた）とエキスをフィルタリング。追加の200μlの分離バッファとメッシュをすすぐ。 </li></ol> 3。胚の分離 3.1スライドの準備<ol> &lt;李&gt;場所倒立顕微鏡のステージ上でクリーン、BSA-コーティングとシリコン処理マルチウェルスライドは、上下に軽くピペッティングによってフィルタリング種子エキスを再懸濁および1または2ウェルに40〜50μlの種子エキスの2滴をピペット。一度に1〜2滴をスクリーニングすると、サンプルの蒸発を防ぎます。 </li><li>前のようにして調製異なるスライドのウェル内の新鮮な分離バッファの代わりに50μlの（1 回目の洗浄の低下）。蒸発を防ぐために覆われ、湿度の高い室内でこのスライドしてください。 </li></ol> 3.2画面、きれい、収集<ol><li> 10倍の倍率の対物レンズで所望の段階で胚のための種子エキスの液滴を選別。必要であれば、20倍の倍率でのステージを確認。胚は通常スライドの底に沈む。 </li><li>手動タングステン針、インスリン注射針または類似の機器と胚の周り破片を除去。 </li><li> T、マイクロマニピュレータを用いた胚の近くにガラスキャピラリーを移動できるだけ少ないソリューションとして持つ胚までAKE。 </li><li>いくつかの胚を（ 例えばすべて1滴）を収集及び第 1 洗浄ドロップ（分子アプリケーション）または宛先バッファ（細胞学的ア ​​プリケーション）でそれらを解放。 （すべて収集ラウンドの総容積は&lt;5μlのでなければなりません）、最小限のボリュームに収集することができるべきである各収集ラウンドは5〜10分であり、胚内に保持されるべきである。 </li><li>胚の所望の量を1 回目の洗浄の低下（中心、可能であれば、記憶を容易にする）で収集されるまで、スクリーニングやコレクションを繰り返します。 </li><li>洗浄降下（破片が回想前に針でそれらを削除し、引き継がれている場合）から、一度にすべての胚を思い出す。 </li><li> 50μlの2 回目の洗浄ドロップに胚を離します。 3.2.6を繰り返します。 </li><li>宛先バッファに胚を離します。転送は、キャピラリー先端の唯一の接触はなく、浸漬し、関与させるべきである。 </li><li> microcapilを交換次のサンプルのためのラリー。 </li></ol>

<ol>
	<li>Gehring, M., Bubb, K. L., Henikoff, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extensive+demethylation+of+repetitive+elements+during+seed+development+underlies+gene+imprinting.">Extensive demethylation of repetitive elements during seed development underlies gene imprinting.</a> Science. 324, 1447-1451 (2009).</li><li>Muller, B., Sheen, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cytokinin+and+auxin+interaction+in+root+stem-cell+specification+during+early+embryogenesis.">Cytokinin and auxin interaction in root stem-cell specification during early embryogenesis.</a> Nature. 453, 1094-1097 (2008).</li><li>Gehring, M., Missirian, V., Henikoff, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genomic+analysis+of+parent-of-origin+allelic+expression+in+Arabidopsis+thaliana+seeds.">Genomic analysis of parent-of-origin allelic expression in Arabidopsis thaliana seeds.</a> PLoS One. 6, 23687-23 (2011).</li><li>Xiang, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genome-wide+analysis+reveals+gene+expression+and+metabolic+network+dynamics+during+embryo+development+in+Arabidopsis.">Genome-wide analysis reveals gene expression and metabolic network dynamics during embryo development in Arabidopsis.</a> Plant Physiol. 156, 346-356 (2011).</li><li>Nawy, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+GATA+factor+HANABA+TARANU+is+required+to+position+the+proembryo+boundary+in+the+early+Arabidopsis+embryo.">The GATA factor HANABA TARANU is required to position the proembryo boundary in the early Arabidopsis embryo.</a> Dev Cell. 19, 103-113 (2010).</li><li>Baroux, C., Pecinka, A., Fuchs, J., Schubert, I., Grossniklaus, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+triploid+endosperm+genome+of+Arabidopsis+adopts+a+peculiar,+parental-dosage-dependent+chromatin+organization.">The triploid endosperm genome of Arabidopsis adopts a peculiar, parental-dosage-dependent chromatin organization.</a> Plant Cell. 19, 1782-1794 (2007).</li><li>Autran, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maternal+epigenetic+pathways+control+parental+contributions+to+Arabidopsis+early+embryogenesis.">Maternal epigenetic pathways control parental contributions to Arabidopsis early embryogenesis.</a> Cell. 145, 707-719 (2011).</li><li>Wohrmann, H. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+a+DNA+methylation-independent+imprinting+control+region+at+the+Arabidopsis+MEDEA+locus.">Identification of a DNA methylation-independent imprinting control region at the Arabidopsis MEDEA locus.</a> Genes Dev. 26, 1837-1850 (2012).</li><li>Raissig, M. T., Baroux, C., Grossniklaus, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regulation+and+Flexibility+of+Genomic+Imprinting+during+Seed+Development.">Regulation and Flexibility of Genomic Imprinting during Seed Development.</a> Plant Cell. 23, 16-26 (2011).</li><li>Rea, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+DNA+methylation+of+imprinted+genes+in+Arabidopsis+endosperm.">Determination of DNA methylation of imprinted genes in Arabidopsis endosperm.</a> J. Vis Exp. 47 (47), e2327 (2011).</li><li>Breuninger, H., Rikirsch, E., Hermann, M., Ueda, M., Laux, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+expression+of+WOX+genes+mediates+apical-basal+axis+formation+in+the+Arabidopsis+embryo.">Differential expression of WOX genes mediates apical-basal axis formation in the Arabidopsis embryo.</a> Dev Cell. 14, 867-876 (2008).</li><li>Kohler, C., Page, D. R., Gagliardini, V., Grossniklaus, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Arabidopsis+thaliana+MEDEA+Polycomb+group+protein+controls+expression+of+PHERES1+by+parental+imprinting.">The Arabidopsis thaliana MEDEA Polycomb group protein controls expression of PHERES1 by parental imprinting.</a> Nature. 37, 28-30 (2005).</li><li>Fransz, P., De Jong, J. H., Lysak, M., Castiglione, M. R., Schubert, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interphase+chromosomes+in+Arabidopsis+are+organized+as+well+defined+chromocenters+from+which+euchromatin+loops+emanate.">Interphase chromosomes in Arabidopsis are organized as well defined chromocenters from which euchromatin loops emanate.</a> Proc. Natl. Acad. Sci. U.S.A. 99, 14584-14589 (2002).</li><li>Morris, J., Singh, J. M., Eberwine, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transcriptome+analysis+of+single+cells.">Transcriptome analysis of single cells.</a> J. Vis. Exp. (50), e2634 (2011).</li><li>Nodine, M. D., Bartel, D. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maternal+and+paternal+genomes+contribute+equally+to+the+transcriptome+of+early+plant+embryos.">Maternal and paternal genomes contribute equally to the transcriptome of early plant embryos.</a> Nature. 482 (7383), (2012).</li><li>Jefferson, R. A., Kavanagh, T. A., Bevan, M. W. G. U. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=fusions:+beta-glucuronidase+as+a+sensitive+and+versatile+gene+fusion+marker+in+higher+plants.">fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants.</a> EMBO J. 6, 3901-3907 (1987).</li><li>Schmidt, A., W&#246;hrmann, H., Raissig, M. T., Arand, J., Gheyselinck, J., Gagliardini, V., Heichinger, C., Walter, J., Grossniklaus, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Polycomb+group+protein+MEDEA+and+the+DNA+methyltransferase+MET1+interact+in+Arabidopsis+to+repress+autonomous+endosperm+development.">The Polycomb group protein MEDEA and the DNA methyltransferase MET1 interact in Arabidopsis to repress autonomous endosperm development.</a> Plant J. 73 (5), (1111).</li></ol>

著者は、彼らが競合する経済的利益を持っていないことを宣言します。

我々は、効果的な、迅速で胚分離プロトコルを開発し、容易に他の研究所に転送することができる。 ここで説明する機器は、倒立顕微鏡、マイクロマニピュレータ、ガラスマイクロキャピラリー、縦フィラメントプラーとマイクロインジェクター（ 図3A）で構成されています。セットアップは、トランスクリプトミクス解析17のための単一の動物細?...

顕花植物の胚は胚乳、第二の受精イベントから派生した栄養組織に囲まれています。胚と胚乳の両方が種皮のいくつかの細胞層に囲まれています。総称して、これらの組織は、果実内部で開発する種子を形成。このように、組織及びシロイヌナズナ胚の細胞特異的な分析が強く、その到達不能が損なわれている。それにもかかわらず、後期球状以降の段階で胚は比較的よく実体顕微鏡下に細いタングステン針を使用して手動で切開の影響を受けやすい、またはそれらを抽出するために鉗子を使用して種子にわずかな圧力を加えることによってです。このような技術は、正常トランスクリプトームまたはそのようなマイクロアレイハイブリダイゼーション、バイサルファイトシークエンシング、又はRNA配列決定（ 例えば 1-3）としてエピゲノムプロファイリング解析に用いた。これとは対照的に、初期の球状ステージへの受精卵における胚の研究は技術的に困難なままです。現在までに、唯一のいくつかの研究では、担当者が持っている固定シードセクション4ま ​​たは罰金ツール5を使用して、種子の中から個々の胚の手動抽出から胚組織のレーザーキャプチャーマイクロダイセクション（LCM）のいずれかを使用して若い胚でortedトランスクリプトーム解析。しかし、LCM装置は早い段階で一般的に利用可能と手動胚抽出ではないことは、時間が容易譲渡することはできません優れた解剖のスキルを消費して必要である。ゲノムワイドな解析に加えて、 その場遺伝子発現解析は、 シロイヌナズナの若者、ホールマウント胚で実行することも困難である。ある程度まで、若い胚は種子穏やかな圧力によって顕微鏡スライド上に解放することができ、（例えば6,7を参照）免疫染色によりレポーター遺伝子アッセイまたはタンパク質検出に使用される。この技術は、しかしながら、このようにして定量分析を妨げ、高スループット胚を分離することはできません。 したがって、我々は、効率的かつ迅速なプロトコルを開発した、簡単に譲渡、下流の様々なアプリケーションに適した設定が簡単であるシロイヌナズナ種子から初期胚の分離のために。適切な分離バッファにプラスチック乳棒を用いてエッペンドルフチュー​​ブの若い長角果から解剖 - 基本的な原理は、優しく、種子を粉砕することです。種子抽出物は、マルチウェルスライド上に液滴内に配置され、倒立顕微鏡を用いて所望の段階で放出胚の存在についてスクリーニングする。胚をマイクロインジェクターまたは標準的な実験室ピペットに装着マイクロキャピラリーガラスを使用して収集されます。分子のアプリケーションでは、胚は最小限のボリュームで宛先バッファに転送する前に、新しい分離バッファの滴に繰り返さリリースで2回洗浄する。細胞学的アプリケーション（レポーターアッセイ、免疫染色、FISH）、洗濯手順については省略することができます。 方法はいくつかの利点を提供します：（i）は、それは細胞学的アプリケーションのための〜45分で25-40の胚が得られlicationsまたは分子アプリケーション用の3-4時間で（洗浄ステップを含む）、（ⅱ）、特定胚の段階の分離を可能にし、（ⅲ）それは、その簡単なセットアップのために他の人や研究室へ簡単に譲渡され、（ⅳ）、それはアップグレードに従順である基本的なセットアップのための手頃な価格の機器が必要であり、および（v）それは正常なRNAシーケン8、遺伝子特異的なDNAメチル化分析9、レポーターアッセイ（10 Raissig らなど、様々なダウンストリームアプリケーションに使用されました）。準備、およびFISH（J.イェーニッシュ、U. Grossniklaus、未発表C. Baroux、 図5を参照）。

<table border="1">
	<tbody>
 <tr>
 <td>Name of the Reagent</td>
 <td>Company</td>
 <td>Catalogue Number</td>
 <td>Comments (optional)</td>
 </tr>
 <tr>
 <td></td>
 <td></td>
 <td></td>
 <td>REAGENTS</td>
 </tr>
 <tr>
 <td>Sigmacote</td>
 <td>SIGMA </td>
 <td>SL2-100 ml</td>
 <td></td>
 </tr>
 <tr>
 <td>RNAse OUT</td>
 <td>Invitrogen (life technologies)</td>
 <td>10777-019</td>
 <td></td>
 </tr>
 <tr>
 <td>First- strand buffer</td>
 <td>Invitrogen (life technologies)</td>
 <td>18064-022</td>
 <td>contained in Superscript II package</td>
 </tr>
 <tr>
 <td>DTT</td>
 <td>Invitrogen (life technologies)</td>
 <td>18064-022</td>
 <td>contained in Superscript II package</td>
 </tr>
 <tr>
 <td>Bovine serum albumin (BSA) 100x =10 mg/ml </td>
 <td>New England Biolabs Inc.</td>
 <td></td>
 <td>Different suppliers will also work </td>
 </tr>
 <tr>
 <td>Thin wall Capillaries 1.0 mm</td>
 <td>World Precision Instruments</td>
 <td>TW100F-4</td>
 <td></td>
 </tr>
 <tr>
 <td>DNA LoBind tube 0.5 ml</td>
 <td>Vaudaux-Eppendorf</td>
 <td>0030108.035</td>
 <td></td>
 </tr>
 <tr>
 <td>CellTrics&Delta; 30 &mu;m</td>
 <td>PARTEC</td>
 <td>04-0042-2316</td>
 <td></td>
 </tr>
 <tr>
 <td>5wells 10 mm diameter slides</td>
 <td>Electron Microscopy Sciences</td>
 <td>63421-10</td>
 <td></td>
 </tr>
 <tr>
 <td>Formaldehyde Solution</td>
 <td>Sigma-Aldrich</td>
 <td>F1635</td>
 <td></td>
 </tr>
 <tr>
 <td>Superfrost Plus slide</td>
 <td>Thermo Fisher</td>
 <td>J1800AMNZ</td>
 <td>Menzel-Gl&auml;ser</td>
 </tr>
 <tr>
 <td>Tris</td>
 <td>Amaresco</td>
 <td>0497</td>
 <td></td>
 </tr>
 <tr>
 <td>EDTA</td>
 <td>Applichem</td>
 <td>A2937</td>
 <td></td>
 </tr>
 <tr>
 <td>Glycin</td>
 <td>Fluka</td>
 <td>50050</td>
 <td></td>
 </tr>
 <tr>
 <td>SDS pellets</td>
 <td>Roth</td>
 <td>CN30.3</td>
 <td></td>
 </tr>
 <tr>
 <td>Micro Pestle</td>
 <td>VWR</td>
 <td>431-0094</td>
 <td></td>
 </tr>
 <tr>
 <td>Microfine insulin syringes</td>
 <td>BD</td>
 <td>U-100</td>
 <td></td>
 </tr>
 <tr>
 <td>DEPC</td>
 <td>Sigma-Aldrich</td>
 <td>D5758</td>
 <td></td>
 </tr>
 <tr>
 <td>Ethanol</td>
 <td>Schaurlau</td>
 <td>ET00102500</td>
 <td></td>
 </tr>
 <tr>
 <td>Forceps N5</td>
 <td>Dumont</td>
 <td>0108-5</td>
 <td></td>
 </tr>
 <tr>
 <td>Bioanalyzer Pico Chip</td>
 <td>Agilent Technologies</td>
 <td>5067-1513</td>
 <td></td>
 </tr>
 <tr>
 <td></td>
 <td></td>
 <td></td>
 <td>EQUIPMENT</td>
 </tr>
 <tr>
 <td>Inverted microscope</td>
 <td>Nikon TMS (Japan),</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>Micromanipulator</td>
 <td>Leitz</td>
 <td></td>
 <td>Leica</td>
 </tr>
 <tr>
 <td>Micomanipulator Post mount LH1 probe</td>
 <td>Leica microsystems</td>
 <td>39430101</td>
 <td>Different brand will also do the work</td>
 </tr>
 <tr>
 <td>Vertical filament puller</td>
 <td>Sutter instrument</td>
 <td>P-20 model</td>
 <td>Other model are also suitable </td>
 </tr>
 <tr>
 <td>Cell Tram vario</td>
 <td>Vaudaux-Eppendorf</td>
 <td>5176.000.033</td>
 <td></td>
 </tr>
 <tr>
 <td>Bioanalyzer</td>
 <td>Agilent Technologies</td>
 <td>2100</td>
 <td></td>
 </tr>
 <tr>
 <td>Qubit Fluorometer</td>
 <td>Invitrogen (life technologies</td>
 <td></td>
 <td></td>
 </tr>
	 </tbody>
 </table>

efficient and rapid isolation of early-stage embryos from arabidopsis thaliana seeds

胚乳 - - 母親の種子の外皮（または種皮）に囲まれた顕花植物では、胚は栄養組織内で開発しています。その結果、初期段階（球状ステージに1セル）での植物の胚の分離が技術的にそれらの相対的な到達不能のために挑戦している。早い段階での効率的な手動の郭清が強く若いシロイヌナズナ種子のサイズが小さく、周囲の組織への胚の接着性によって損なわれている。ここでは、下流のアプリケーションに応じて、1時間〜4時間で40胚まで降伏、若いシロイヌナズナ胚の効率的な分離を可能にする方法について説明します。胚はわずかにエッペンドルフチュー​​ブ内でプラスチック乳棒で250から750種を粉砕することにより単離緩衝液中に放出される。ガラスピペット（ゴムチューブを介して）または油圧制御マイクロインジェクターを滴場所から胚を収集するために使用される標準的な実験室のどちらかに接続されているマイクロキャピラリー倒立光学顕微鏡にマルチウェルスライド上のD。必要な技術的なスキルは、シンプルかつ簡単に譲渡され、基本的なセットアップは、高価な機器を必要としません。収集された胚は、例えばRT-PCR、RNA配列決定、DNAメチル化分析、in situハイブリダイゼーション（FISH） の蛍光、免疫染色、およびレポーター遺伝子アッセイなどの下流の種々の用途に適している。

洗浄、分子のアプリケーションに、または洗浄の細胞学的ア ​​プリケーションのために、たとえば 、省略されている場合、時間足らずで例えば 、実行している場合は私たちの胚の分離手順（ 図1）は、4時間で40胚までの分離を可能にします。2ディスプレイ高低図品質マイクロキャピラリー先端と図3は胚分離機のセットアッ?...

Watch this Scientific Journal Video about Efficient and Rapid Isolation of Early-stage Embryos from Arabidopsis thaliana Seeds at JoVE.com

Efficient and Rapid Isolation of Early-stage Embryos from Arabidopsis thaliana Seeds

我々は、より開発の初期段階で胚を単離するための効率的で簡単な方法を報告する<em&gt;シロイヌナズナ</em&gt;種。最大40胚には、下流のアプリケーションに応じて、1時間〜4時間で単離することができる。手順は、DNAのメチル化は、レポーター遺伝子の発現、および免疫蛍光トランスクリプトームのために適している<em&gt;その場で</em&gt;ハイブリダイゼーション分析。

から初期段階の胚の効率的かつ迅速な単離<em&gt;シロイヌナズナ</em&gt;種子

In flowering plants, the embryo develops within a nourishing tissue - the endosperm - surrounded by the maternal seed integuments (or seed coat). As a ...

efficient-rapid-isolation-early-stage-embryos-from-arabidopsis

Research

JoVE Journal

Biology

Efficient and Rapid Isolation of Early-stage Embryos from Arabidopsis thaliana Seeds

17.7K ビュー.  University of Zürich. 我々は、より開発の初期段階で胚を単離するための効率的で簡単な方法を報告するシロイヌナズナ種。最大40胚には、下流のアプリケーションに応じて、1時間〜4時間で単離することができる。手順は、DNAのメチル化は、レポーター遺伝子の発現、および免疫蛍光トランスクリプトームのために適しているその場でハイブリダイゼー?...

ビデオ: Efficient and Rapid Isolation of Early-stage Embryos from Arabidopsis thaliana Seeds

Placing Growth Factor-Coated Beads on Early Stage Chicken Embryos

The neural tube expresses many proteins in specific spatiotemporal patterns during development. These proteins have been shown to be critical for cell fate determination, cell migration, and formation of neural circuits. Neuronal induction and patterning involve bone morphogenetic protein (BMP), sonic hedgehog (SHH), fibroblast growth factor (FGF), among others. In particular, the expression pattern of Fgf8 is in close proximity to regions expressing BMP4 and SHH. This expression pattern is consistent with developmental interactions that facilitate patterning in the telencephalon.

Here we provide a visual demonstration of a method in which an in ovo preparation can be used to test the effects of Fgfs in the formation of the forebrain. Beads are coated with protein and placed in the developing neural tube to provide sustained exposure. Because the procedure uses small, carefully placed beads, it is minimally invasive and allows several beads to be placed within a single neural tube. Moreover, the method allows for continued development so that embryos can be analyzed at a more mature stage to detect changes in anatomy and in neural patterning. This simple but useful protocol allows for real time imaging. It provides a means to make spatially and temporally limited changes to endogenous protein levels.

A variety of growth factors and proteins interact to induce cells to take on different cell fates during development. Here we demonstrate the use of an in ovo preparation to address possible interactions between different proteins in development by placing beads on E2.5 chick embryos.

アーリーステージのニワトリ胚に成長因子コートビーズの配置

The neural tube expresses many proteins in specific spatiotemporal patterns during development.   These proteins have been shown to be critical for cell ...

生物学

Isolation of Protoplasts from Tissues of 14-day-old Seedlings of Arabidopsis thaliana

Protoplasts are plant cells that have had their cell walls enzymatically removed. Isolation of protoplasts from different plant tissues was first reported more than 40 years ago 1 and has since been adapted to study a variety of cellular processes, such as subcellular localization of proteins, isolation of intact organelles and targeted gene-inactivation by double stranded RNA interference (RNAi) 2-5. Most of the protoplast isolation protocols use leaf tissues of mature Arabidopsis (e.g. 35-day-old plants) 2-4. We modified existing protocols by employing 14-day-old Arabidopsis seedlings. In this procedure, one gram of 14-day-old seedlings yielded 5 106-107 protoplasts that remain intact at least 96 hours. The yield of protoplasts from seedlings is comparable with preparations from leaves of mature Arabidopsis, but instead of 35-36 days, isolation of protoplasts is completed in 15 days. This allows decreasing the time and growth chamber space that are required for isolating protoplasts when mature plants are used, and expedites the downstream studies that require intact protoplasts.

Isolation of Protoplasts from Tissues of 14-day-old Seedlings of Arabidopsis thaliana

This video shows a procedure for isolating intact protoplasts from tissues of 14-day-old seedlings of Arabidopsis. Given that the isolated protoplasts remain intact for at least 96h and are isolated from seedlings instead of one-month-old mature plants, this procedure expedites assays requiring intact protoplasts.

の14日齢の苗の組織からのプロトプラストの単離シロイヌナズナ</em

Protoplasts are plant cells that have had their cell walls enzymatically removed. Isolation of protoplasts  from different plant tissues was first ...

Floral-dip Transformation of Arabidopsis thaliana to Examine pTSO2::&beta;-glucuronidase Reporter Gene Expression

The ability to introduce foreign genes into an organism is the foundation for modern biology and biotechnology. In the model flowering plant Arabidopsis thaliana, the floral-dip transformation method1-2 has replaced all previous methods because of its simplicity, efficiency, and low cost. Specifically, shoots of young flowering Arabidopsis plants are dipped in a solution of Agrobacterium tumefaciens carrying specific plasmid constructs. After dipping, the plants are returned to normal growth and yield seeds, a small percentage of which are transformed with the foreign gene and can be selected for on medium containing antibiotics. This floral-dip method significantly facilitated Arabidopsis research and contributed greatly to our understanding of plant gene function. In this study, we use the floral-dip method to transform a reporter gene, &beta;-glucuronidase (GUS), under the control of TSO2 promoter. TSO2, coding for the Ribonucleotide Reductase (RNR) small subunit3, is a cell cycle regulated gene essential for dNDP biosynthesis in the S-phase of the cell cycle. Examination of GUS expression in transgenic Arabidopsis seedlings shows that TSO2 is expressed in actively dividing tissues. The reported experimental method and materials can be easily adapted not only for research but also for education at high school and college levels.

Floral-dip Transformation of Arabidopsis thaliana to Examine pTSO2::&#946;-glucuronidase Reporter Gene Expression

This article illustrates the floral-dip method of Agrobacterium tumefaciens -mediated transformation of Arabidopsis thaliana. By introducing a cell-cycle regulated promoter-reporter, pTSO2::&beta;-glucuronidase (GUS), into Arabidopsis, we illustrates how one detects GUS reporter expression in transgenic seedlings.

フローラルディップ変換のシロイヌナズナ調べに pTSO2：：β-グルクロニダーゼレポーター遺伝子発現

The ability to introduce foreign genes into an organism is the foundation for modern biology and biotechnology. In the model flowering plant Arabidopsis ...

Arabidopsis thaliana Polar Glycerolipid Profiling by Thin Layer Chromatography (TLC) Coupled with Gas-Liquid Chromatography (GLC)

Biological membranes separate cells from the environment. From a single cell to multicellular plants and animals, glycerolipids, such as phosphatidylcholine or phosphatidylethanolamine, form bilayer membranes which act as both boundaries and interfaces for chemical exchange between cells and their surroundings. Unlike animals, plant cells have a special organelle for photosynthesis, the chloroplast. The intricate membrane system of the chloroplast contains unique glycerolipids, namely glycolipids lacking phosphorus: monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), sulfoquinovosyldiacylglycerol (SQDG)4. The roles of these lipids are beyond simply structural. These glycolipids and other glycerolipids were found in the crystal structures of photosystem I and II indicating the involvement of glycerolipids in photosynthesis8,11. During phosphate starvation, DGDG is transferred to extraplastidic membranes to compensate the loss of phospholipids9,12.
Much of our knowledge of the biosynthesis and function of these lipids has been derived from a combination of genetic and biochemical studies with Arabidopsis thaliana14. During these studies, a simple procedure for the analysis of polar lipids has been essential for the screening and analysis of lipid mutants and will be outlined in detail. A leaf lipid extract is first separated by thin layer chromatography (TLC) and glycerolipids are stained reversibly with iodine vapor. The individual lipids are scraped from the TLC plate and converted to fatty acyl methylesters (FAMEs), which are analyzed by gas-liquid chromatography coupled with flame ionization detection (FID-GLC) (Figure 1). This method has been proven to be a reliable tool for mutant screening. For example, the tgd1,2,3,4 endoplasmic reticulum-to-plastid lipid trafficking mutants were discovered based on the accumulation of an abnormal galactoglycerolipid: trigalactosyldiacylglycerol (TGDG) and a decrease in the relative amount of 18:3 (carbons : double bonds) fatty acyl groups in membrane lipids 3,13,18,20. This method is also applicable for determining enzymatic activities of proteins using lipids as substrate6.

Arabidopsis thaliana Polar Glycerolipid Profiling by Thin Layer Chromatography TLC Coupled with Gas-Liquid Chromatography GLC

Composition of polar lipid extracts and the fatty acid composition of individual glycerolipids are determined in a simple and robust lipid profiling experiment. For this purpose, glycerolipids are isolated by thin layer chromatography and subjected to transmethylation of their acyl groups. Fatty acyl methylesters are quantified by gas-liquid chromatography.

気-液クロマトグラフィー（GLC）とあいまって薄層クロマトグラフィー（TLC）によりシロイヌナズナポーラーグリセロ脂質プロファイリング

Biological membranes separate cells from the environment. From a single cell to multicellular plants and animals, glycerolipids, such as ...

Chromatin Immunoprecipitation Assay for Tissue-specific Genes using Early-stage Mouse Embryos

Chromatin immunoprecipitation (ChIP) is a powerful tool to identify protein:chromatin interactions that occur in the context of living cells 1-3. This technique has been widely exploited in tissue culture cells, and to a lesser extent, in primary tissue. The application of ChIP to rodent embryonic tissue, especially at early times of development, is complicated by the limited amount of tissue and the heterogeneity of cell and tissue types in the embryo. Here we present a method to perform ChIP using a dissociated embryonic day 8.5 (E8.5) embryo. Sheared chromatin from a single E8.5 embryo can be divided into up to five aliquots, which allows the investigator sufficient material for controls and for investigation of specific protein:chromatin interactions.
We have utilized this technique to begin to document protein:chromatin interactions during the specification of tissue-specific gene expression programs. The heterogeneity of cell types in an embryo necessarily restricts the application of this technique because the result is the detection of protein:chromatin interactions without distinguishing whether the interactions occur in all, a subset of, or a single cell type(s). However, examination of tissue-specific genes during or following the onset of tissue-specific gene expression is feasible for two reasons. First, immunoprecipitation of tissue specific factors necessarily isolates chromatin from the cell type where the factor is expressed. Second, immunoprecipitation of coactivators and histones containing post-translational modifications that are associated with gene activation should only be found at genes and gene regulatory sequences in the cell type where the gene is being or has been activated. The technique should be applicable to the study of most tissue-specific gene activation events.
In the example described below, we utilized E8.5 and E9.5 mouse embryos to examine factor binding at a skeletal muscle specific gene promoter. Somites, which are the precursor tissues from which the skeletal muscles of the trunk and limbs will form, are present at E8.5-9.54,5. Myogenin is a regulatory factor required for skeletal muscle differentiation 6-9. The data demonstrate that myogenin is associated with its own promoter in E8.5 and E9.5 embryos. Because myogenin is only expressed in somites at this stage of development 6,10, the data indicate that myogenin interactions with its own promoter have already occurred in skeletal muscle precursor cells in E8.5 embryos.

We demonstrate a chromatin immunoprecipitation (ChIP) method to identify factor interactions at tissue-specific genes during or after the onset of tissue-specific gene expression in mouse embryonic tissue. This protocol should be widely applicable for the study of tissue-specific gene activation as it occurs during normal embryonic development.

初期段階のマウス胚を用いて組織特異的遺伝子のためのクロマチン免疫沈降アッセイ

Chromatin immunoprecipitation (ChIP) is a powerful tool to identify protein:chromatin interactions that occur in the context of living cells 1-3. This ...

Isolation and Purification of Kinesin from Drosophila Embryos

Motor proteins move cargos along microtubules, and transport them to specific sub-cellular locations. Because altered transport is suggested to underlie a variety of neurodegenerative diseases, understanding microtubule based motor transport and its regulation will likely ultimately lead to improved therapeutic approaches. Kinesin-1 is a eukaryotic motor protein which moves in an anterograde (plus-end) direction along microtubules (MTs), powered by ATP hydrolysis. Here we report a detailed purification protocol to isolate active full length kinesin from Drosophila embryos, thus allowing the combination of Drosophila genetics with single-molecule biophysical studies. Starting with approximately 50 laying cups, with approximately 1000 females per cup, we carried out overnight collections. This provided approximately 10 ml of packed embryos. The embryos were bleach dechorionated (yielding approximately 9 grams of embryos), and then homogenized. After disruption, the homogenate was clarified using a low speed spin followed by a high speed centrifugation. The clarified supernatant was treated with GTP and taxol to polymerize MTs. Kinesin was immobilized on polymerized MTs by adding the ATP analog, 5'-adenylyl imidodiphosphate at room temperature. After kinesin binding, microtubules were sedimented via high speed centrifugation through a sucrose cushion. The microtubule pellet was then re-suspended, and this process was repeated. Finally, ATP was added to release the kinesin from the MTs. High speed centrifugation then spun down the MTs, leaving the kinesin in the supernatant. This kinesin was subjected to a centrifugal filtration using a 100 KD cut off filter for further purification, aliquoted, snap frozen in liquid nitrogen, and stored at -80 &deg;C. SDS gel electrophoresis and western blotting was performed using the purified sample. The motor activity of purified samples before and after the final centrifugal filtration step was evaluated using an in vitro single molecule microtubule assay. The kinesin fractions before and after the centrifugal filtration showed processivity as previously reported in literature. Further experiments are underway to evaluate the interaction between kinesin and other transport related proteins.

Isolation and Purification of Kinesin from Drosophila Embryos

This is a protocol to isolate active full length Kinesin from Drosophila embryos for single-molecule biophysical studies. We show how to collect embryos, make the embryo lysate, and then polymerize microtubules (MTs). Kinesin is purified by immobilizing it on the MTs, spinning down the Kinesin-MT complexes, and then releasing the kinesin from the MTs via ATP addition.

キネシンの単離及び精製からショウジョウバエ胚

Motor proteins move cargos along microtubules, and transport them to specific sub-cellular locations. Because altered transport is suggested to underlie a ...

Rapid and Efficient Generation of Neurons from Human Pluripotent Stem Cells in a Multititre Plate Format

Existing protocols for the generation of neurons from human pluripotent stem cells (hPSCs) are often tedious in that they are multistep procedures involving the isolation and expansion of neural precursor cells, prior to terminal differentiation. In comparison to these time-consuming approaches, we have recently found that combined inhibition of three signaling pathways, TGF&beta;/SMAD2, BMP/SMAD1, and FGF/ERK, promotes rapid induction of neuroectoderm from hPSCs, followed by immediate differentiation into functional neurons. Here, we have adapted our procedure to a novel multititre plate format, to further enhance its reproducibility and to make it compatible with mid-throughput applications. It comprises four days of neuroectoderm formation in floating spheres (embryoid bodies), followed by a further four days of differentiation into neurons under adherent conditions. Most cells obtained with this protocol appear to be bipolar sensory neurons. Moreover, the procedure is highly efficient, does not require particular expert skills, and is based on a simple chemically defined medium with cost-efficient small molecules. Due to these features, the procedure may serve as a useful platform for further functional investigation as well as for cell-based screening approaches requiring human sensory neurons or neurons of any type.

Protocols for neuronal differentiation of pluripotent human stem cells (hPSCs) are often time-consuming and require substantial cell culture skills. Here, we have adapted a small molecule-based differentiation procedure to a multititre plate format, allowing simple, rapid, and efficient generation of human neurons in a controlled manner.

Multititreプレートフォーマットでヒト多能性幹細胞から神経細胞の迅速かつ効率的な生成

Existing protocols for the generation of neurons from human pluripotent stem cells (hPSCs) are often tedious in that they are  multistep procedures ...

Metabolic Labeling and Membrane Fractionation for Comparative Proteomic Analysis of Arabidopsis thaliana Suspension Cell Cultures

Plasma membrane microdomains are features based on the physical properties of the lipid and sterol environment and have particular roles in signaling processes. Extracting sterol-enriched membrane microdomains from plant cells for proteomic analysis is a difficult task mainly due to multiple preparation steps and sources for contaminations from other cellular compartments. The plasma membrane constitutes only about 5-20% of all the membranes in a plant cell, and therefore isolation of highly purified plasma membrane fraction is challenging. A frequently used method involves aqueous two-phase partitioning in polyethylene glycol and dextran, which yields plasma membrane vesicles with a purity of 95% 1. Sterol-rich membrane microdomains within the plasma membrane are insoluble upon treatment with cold nonionic detergents at alkaline pH. This detergent-resistant membrane fraction can be separated from the bulk plasma membrane by ultracentrifugation in a sucrose gradient 2. Subsequently, proteins can be extracted from the low density band of the sucrose gradient by methanol/chloroform precipitation. Extracted protein will then be trypsin digested, desalted and finally analyzed by LC-MS/MS. Our extraction protocol for sterol-rich microdomains is optimized for the preparation of clean detergent-resistant membrane fractions from Arabidopsis thaliana cell cultures.
We use full metabolic labeling of Arabidopsis thaliana suspension cell cultures with K15NO3 as the only nitrogen source for quantitative comparative proteomic studies following biological treatment of interest 3. By mixing equal ratios of labeled and unlabeled cell cultures for joint protein extraction the influence of preparation steps on final quantitative result is kept at a minimum. Also loss of material during extraction will affect both control and treatment samples in the same way, and therefore the ratio of light and heave peptide will remain constant. In the proposed method either labeled or unlabeled cell culture undergoes a biological treatment, while the other serves as control 4.

Metabolic Labeling and Membrane Fractionation for Comparative Proteomic Analysis of Arabidopsis thaliana Suspension Cell Cultures

Here we describe a robust method for the fractionation of plant plasma membranes into detergent resistant and detergent soluble membranes based on a mixture of unlabeled and in vivo fully 15N labeled Arabidopsis thaliana cell cultures. The procedure is applied for comparative proteomic studies to understand signaling processes.

比較プロテオーム解析のための代謝標識および膜分画<em&gt;シロイヌナズナ</em&gt;サスペンション細胞培養

Plasma membrane microdomains are features based on the physical properties of the lipid and sterol environment and have particular roles in signaling ...

Histochemical Staining of Arabidopsis thaliana Secondary Cell Wall Elements

Arabidopsis thaliana is a model organism commonly used to understand and manipulate various cellular processes in plants, and it has been used extensively in the study of secondary cell wall formation. Secondary cell wall deposition occurs after the primary cell wall is laid down, a process carried out exclusively by specialized cells such as those forming vessel and fiber tissues. Most secondary cell walls are composed of cellulose (40&#8211;50%), hemicellulose (25&#8211;30%), and lignin (20&#8211;30%). Several mutations affecting secondary cell wall biosynthesis have been isolated, and the corresponding mutants may or may not exhibit obvious biochemical composition changes or visual phenotypes since these mutations could be masked by compensatory responses. Staining procedures have historically been used to show differences on a cellular basis. These methods are exclusively visual means of analysis; nevertheless their role in rapid and critical analysis is of great importance. Congo red and calcofluor white are stains used to detect polysaccharides, whereas M&#228;ule and phloroglucinol are commonly used to determine differences in lignin, and toluidine blue O is used to differentially stain polysaccharides and lignin. The seemingly simple techniques of sectioning, staining, and imaging can be a challenge for beginners. Starting with sample preparation using the A. thaliana model, this study details the protocols of a variety of staining methodologies that can be easily implemented for observation of cell and tissue organization in secondary cell walls of plants.

Histochemical Staining of Arabidopsis thaliana Secondary Cell Wall Elements

Plant cell wall composition varies between tissue types and can include lignin, cellulose, hemicelluloses, and pectin. Various staining techniques have been developed to visualize differences at the cell-type level. This paper is a compilation of commonly used cell wall staining techniques.

の組織化学的染色<em&gt;シロイヌナズナ</em&gt;二次細胞壁の要素

Arabidopsis thaliana is a model organism commonly used to understand and manipulate various cellular processes in plants, and it has been used extensively ...

Standardized Method for High-throughput Sterilization of Arabidopsis Seeds

Arabidopsis thaliana (Arabidopsis) seedlings often need to be grown on sterile media. This requires prior seed sterilization to prevent the growth of microbial contaminants present on the seed surface. Currently, Arabidopsis seeds are sterilized using two distinct sterilization techniques in conditions that differ slightly between labs and have not been standardized, often resulting in only partially effective sterilization or in excessive seed mortality. Most of these methods are also not easily scalable to a large number of seed lines of diverse genotypes. As technologies for high-throughput analysis of Arabidopsis continue to proliferate, standardized techniques for sterilizing large numbers of seeds of different genotypes are becoming essential for conducting these types of experiments. The response of a number of Arabidopsis lines to two different sterilization techniques was evaluated based on seed germination rate and the level of seed contamination with microbes and other pathogens. The treatments included different concentrations of sterilizing agents and times of exposure, combined to determine optimal conditions for Arabidopsis seed sterilization. Optimized protocols have been developed for two different sterilization methods: bleach (liquid-phase) and chlorine (Cl2) gas (vapor-phase), both resulting in high seed germination rates and minimal microbial contamination. The utility of these protocols was illustrated through the testing of both wild type and mutant seeds with a range of germination potentials. Our results show that seeds can be effectively sterilized using either method without excessive seed mortality, although detrimental effects of sterilization were observed for seeds with lower than optimal germination potential. In addition, an equation was developed to enable researchers to apply the standardized chlorine gas sterilization conditions to airtight containers of different sizes. The protocols described here allow easy, efficient, and inexpensive seed sterilization for a large number of Arabidopsis lines.

The objective of this study was to determine the effects of bleach and chlorine gas sterilization on seed germination of a range of Arabidopsis genotypes grown on sterile media. Optimized sterilization protocols have been developed to prevent the growth of microbial contaminants while providing satisfactory seed survival.

高スループット シロイヌナズナ種子の殺菌のための標準化された方法

Arabidopsis thaliana (Arabidopsis) seedlings often need to be grown on sterile media. This requires prior seed sterilization to prevent the growth of ...

から初期段階の胚の効率的かつ迅速な単離

要約

さらに動画を探す

この動画の章

アーリーステージのニワトリ胚に成長因子コートビーズの配置

の14日齢の苗の組織からのプロトプラストの単離シロイヌナズナ

フローラルディップ変換のシロイヌナズナ調べに pTSO2：：β-グルクロニダーゼレポーター遺伝子発現

気-液クロマトグラフィー（GLC）とあいまって薄層クロマトグラフィー（TLC）によりシロイヌナズナポーラーグリセロ脂質プロファイリング

初期段階のマウス胚を用いて組織特異的遺伝子のためのクロマチン免疫沈降アッセイ

キネシンの単離及び精製からショウジョウバエ胚

Multititreプレートフォーマットでヒト多能性幹細胞から神経細胞の迅速かつ効率的な生成

比較プロテオーム解析のための代謝標識および膜分画

の組織化学的染色

高スループットシロイヌナズナ種子の殺菌のための標準化された方法

から初期段階の胚の効率的かつ迅速な単離

要約

さらに動画を探す

この動画の章

アーリーステージのニワトリ胚に成長因子コートビーズの配置

の14日齢の苗の組織からのプロトプラストの単離シロイヌナズナ

フローラルディップ変換のシロイヌナズナ調べに pTSO2：：β-グルクロニダーゼレポーター遺伝子発現

気-液クロマトグラフィー（GLC）とあいまって薄層クロマトグラフィー（TLC）によりシロイヌナズナポーラーグリセロ脂質プロファイリング

初期段階のマウス胚を用いて組織特異的遺伝子のためのクロマチン免疫沈降アッセイ

キネシンの単離及び精製からショウジョウバエ胚

Multititreプレートフォーマットでヒト多能性幹細胞から神経細胞の迅速かつ効率的な生成

比較プロテオーム解析のための代謝標識および膜分画

の組織化学的染色

高スループット シロイヌナズナ種子の殺菌のための標準化された方法

高スループットシロイヌナズナ種子の殺菌のための標準化された方法