Name: 内在コントラストを用いたDNA分子の分光超分解能イメージング (動画) | JoVE Journal
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Description: 学ぶ 分光単分子局在顕微鏡を用いたDNA分子のラベルフリー超分解能イメージング手法。内在蛍光を活用し、外部標識なしでDNAのネイティブ光学特性と配列情報を保持しながら高分解能イメージングを実現します。

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</ol>

<table><tbody><tr><td>イメージング用488 nm発光フィルター</td><td>クロマ・テクノロジー社</td><td>ET525/50M</td><td></td></tr><tr><td>校正用の532 nmバンドパスフィルター</td><td>ソーラブ</td><td>FL532-3</td><td></td></tr><tr><td>イメージング用の532 nm放出フィルター</td><td>クロマ・テクノロジー社</td><td>ET620/60M</td><td></td></tr><tr><td>580 nmバンドパスフィルターによる校正</td><td>ソーラブ</td><td>FB580-10</td><td></td></tr><tr><td>キャリブレーション用の633 nmバンドパスフィルター</td><td>ソーラブ</td><td>FL632.8-3</td><td></td></tr><tr><td>校正用の680 nmバンドパスフィルタ</td><td>ソーラブ</td><td>FB680-10</td><td></td></tr><tr><td>キャリブレーション用の750 nmバンドパスフィルター</td><td>ソーラブ</td><td>FB750-10</td><td></td></tr><tr><td>ベータ-メルカプトエタノール&nbsp;</td><td>シグマ</td><td>M6250</td><td></td></tr><tr><td>BMEイメージングバッファ</td><td>NA</td><td>NA</td><td>7 & mu;L・グロックス、7 & mu;Lベータメルカプトエタノールおよび690 & mu;Lグルコースバッファー</td></tr><tr><td>牛肝臓由来のカタラーゼ</td><td>シグマ</td><td>C100</td><td></td></tr><tr><td>DABCO(1,4-ジアゾビシクロ-(2.2.2)-オクタン)</td><td>シグマ</td><td>D27802</td><td></td></tr><tr><td>DABCOイメージングバッファ</td><td>NA</td><td>NA</td><td>65 mM DABCO+30 mM 硫酸ナトリウム + 30 mM DTT 1 M&nbsp;DNAseではRNAseフリーの脱イオン水が使われています。亜硫酸ナトリウムはPBSに10&10倍溶けるべきです。イメージングバッファを作成する前に1 Mまで。</td></tr><tr><td>分光SMLMのためのDNAデプレックス</td><td>IDT</td><td>NA</td><td>20塩基対の二本鎖GC_alter、GC_conti、AT_alter、AT_conti. 例:GC_alterは両方の鎖を5'としています- GCGCGCGGGGGGCGCC - 3フィート;GC_conti 1本のストランドが5フィートです- ググググ - 3フィートもう1つは5フィート- CCCCCCCCCCCCCCCCCCCC - 3' すべての試料はHPLC規格です。</td></tr><tr><td>DNAオリゴヌクレオチド/二本鎖DNA</td><td>IDT</td><td>NA</td><td></td></tr><tr><td>分光SMLMにおけるDNAオリゴ</td><td>IDT</td><td>NA</td><td>20基座A、G、C、T;40基底単一鎖交流チェーン(A1C1、A5C5、A10C10、A20C20);&nbsp;単鎖5、8、12、16塩基グアニン。 例:A10C10は 5'- AAAAAAAAACCCC CCCCCAAAAAAAAAAAA AACCCCCCCC - 3' すべてのサンプルはHPLC規格です。</td></tr><tr><td>DNase/RNaseフリー蒸留水</td><td>インビトロジェン</td><td>10977015</td><td></td></tr><tr><td>DTT(DL-ジチオスライトール)</td><td>シグマ</td><td>43816</td><td></td></tr><tr><td>グロックス</td><td>NA</td><td>NA</td><td>14 mgグルコースオキシダーゼ、50 & Mu;Lカタラーゼ(17 mg/ml)+200 & μ;LバッファーAは10 mMトリス(pH 8.0)、50 mM NaCl.&nbsp;</td></tr><tr><td>グルコース緩衝剤</td><td>NA</td><td>NA</td><td>50 mM Tris(pH 8.0)、50 mM NaCl、10%グルコース & nbsp;</td></tr><tr><td>グルコース酸化酵素</td><td>シグマ</td><td>G7141</td><td></td></tr><tr><td>グルコース、粉末</td><td>サーモ・フィッシャー</td><td>15023021</td><td></td></tr><tr><td>PBS(10X)</td><td>サーモ・フィッシャー</td><td>J62036。K3</td><td></td></tr><tr><td>PBS(1X)</td><td>ギブコ</td><td>10010023</td><td></td></tr><tr><td>塩化ナトリウム</td><td>サーモ・フィッシャー</td><td>424290010</td><td></td></tr><tr><td>亜硫酸ナトリウム&nbsp;</td><td>シグマ</td><td>S0505</td><td></td></tr><tr><td>トリス(1M) pH 8.0</td><td>インビトロジェン</td><td>AM9856</td><td></td></tr></tbody></table>

内在コントラストを用いたdna分子の分光超分解能イメージング

Spectroscopic Super-resolution Imaging of DNA Molecules using Intrinsic Contrast

内在コントラストを用いたDNA分子の分光超分解能イメージング

Super-resolution imaging has revolutionized biological research by revealing structural details at the nanoscale. Most optical super-resolution methods ...

spectroscopic-super-resolution-imaging-dna-molecules-using-intrinsic

Research

JoVE Journal

Bioengineering

バイオエンジニアリング（生体工学）

この動画では、特定の遺伝子座におけるDNA分子上の融合タンパク質凝縮物の集合を、1分子イメージングを用いて可視化する方法を説明しています。これらの凝縮物は、LCD-LCDおよびLCD-DBDの相互作用によって形成され、RNAポリメラーゼIIの動員と遺伝子転写の促進を可能にします。...

DNAカーテン上の融合タンパク質凝縮物の集合を可視化するための1分子イメージング

我々は、細菌のFtsZ-リングの構造、組織、細胞分裂に不可欠な装置を検査する、超解像イメージング法を説明します。この方法は、光活​​性化ローカライゼーション顕微鏡（PALM）画像の定量分析に基づいており、他の細菌の細胞骨格タンパク質に適用することができます。...

細菌分裂機械の超解像イメージング

新たに発見された 5-メチルシトシン (oxi mCs), 5-ヒドロキシメチルシトシン (5hmC), 5-formylcytosine (5 fC) の酸化フォームと 5-carboxylcytosine (5caC) は、独自の機能的役割の異なる DNA 変更を表す場合があります。ここの可視化から oxi mCs...

DNA 変更染色: 共焦点画像の数値解析

この方法論の全体的な目標は、固定細胞における微小管、中間径フィラメントの 2D デュアル カラー dSTORM...

中間径フィラメントと微小管 2次元直接確率光再構成顕微鏡イメージング

詳細な指示は、タイル (HIST) 顕微鏡と 1...

高傾斜の構築ガイド拡張ビュー フィールドの単一分子イメージング用タイル顕微鏡を掃引

ヒト初等細胞の核アーキテクチャを研究する3D多色DNA FISHツール

本プロトコルは、単一分子局在顕微鏡データ、既知の体積、ゲノムサイズ、および細胞周期段階のボロノイテッセレーションを使用して、接着細胞核内の絶対DNA密度を測定する方法を説明する。...

単一分子局在顕微鏡を用いた細胞核中の絶対DNA密度のマッピング

このプロトコルは、CMGヘリカーゼによるDNA巻き戻しのライブ可視化のための単一分子アッセイの実施を示しています。(1)DNA基質の調製、(2)蛍光標識 されたDrosophila melanogaster...

CMGヘリカーゼによるDNA巻き戻しの1分子リアルタイム可視化

このプロトコルは、光学光子再割り当てを備えた回転円盤共焦点顕微鏡を用いて深部全細胞単分子局在顕微鏡(SMLM)を行う手順を示しており、カスタム光学や複雑な照明を必要とせずにDNA-PAINTイメージングを可能にします。...

光学光子再割り当て を用いた回転円 板共焦点によるDNA-PAINTによる全セル超分解能イメージング

核酸、タンパク質と低分子のReagentless検出用電気化学DNAバイオセンサーの作製

学ぶ 分光単分子局在顕微鏡を用いたDNA分子のラベルフリー超分解能イメージング手法。内在蛍光を活用し、外部標識なしでDNAのネイティブ光学特性と配列情報を保持しながら高分解能イメージングを実現します。

内在コントラストを用いたDNA分子の分光超分解能イメージング

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