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[FLAG®および3xFLAG™エピトープに対する高特異性抗体](https://www.sigmaaldrich.com#flag5) - [SnapFast™ FLAG®および3xFLAG™ベクター従来製品比較表](https://www.sigmaaldrich.com#flag6) ## [](https://www.sigmaaldrich.com)FLAG®タグ発現ベクターマップおよび3xFLAG™ペプチド 細菌および哺乳類の発現ベクター（もしくは発現コンストラクト）は、目的の組換えタンパク質を過剰発現させるために一般的に使用される、環状DNAトランスフェクション用プラスミドです。ベクターコンストラクト内にエンコードされた固有の配列は、トランスフェクトされた細胞自体のタンパク質合成機構によって認識されるプロモーター要素を含んでおり、取り込まれたタンパク質配列の発現を促進します。 FLAG®発現システムは、組換え融合タンパク質を発現、精製、検出用に確立されたシステムです。FLAG®および3xFLAG™試薬は、ウェスタンブロッティング、免疫細胞化学、免疫沈降、フローサイトメトリー、タンパク質精製のほか、タンパク質間相互作用、細胞の微細構造、タンパク質同定の研究など、さまざまな用途に有用であることが証明されています。このスモールサイズの親水性タグを使うことにより、特異性と感度の高いANTI-FLAG®抗体での組換え融合タンパク質の検出と精製がさらに容易になります。 ## [](https://www.sigmaaldrich.com)FLAG®および3xFLAG™エピトープタグDNA配列／アミノ酸ペプチド配列 FLAG®発現システムは、目的の組換えタンパク質に融合された短い親水性8アミノ酸ペプチドを使用します。FLAG®ペプチドは親水性のため、融合タンパク質の表面付近に存在する傾向があります。そのため、エンテロキナーゼ（Ek）によるタグの切断と抗体による検出も容易になります。また、FLAG®ペプチドタグのサイズは小さいため、他のエピトープやドメインを覆ってしまったり、融合タンパク質の機能や分泌、移動を変えてしまう可能性がほとんどありません。 3xFLAG™システムは、3つのタンデムFLAG®エピトープ（22アミノ酸）を融合させた改良システムです。3xFLAG™を含む融合タンパク質の検出感度は、他のシステムを最大で200倍上回ります。従来のFLAG®タグと同様に、3xFLAG™は親水性で、エンテロキナーゼ切断部位を有し、比較的小さいため、タンパク質機能の変化、他のエピトープタグのブロック、溶解性の低下等の可能性が最小限に抑えられています。  #### FLAG®および3xFLAG™タグのアミノ酸配列 エンテロキナーゼを使用すると、タグのC末端にあるAsp-Asp-Asp-Asp-Lysアミノ酸配列の下流で切断することにより、N末端FLAG®（8アミノ酸）および3xFLAG™（22アミノ酸）タグの除去が可能となります。 ## [](https://www.sigmaaldrich.com)FLAG®および3xFLAG™細菌／哺乳類発現ベクター製品 従来のFLAG®および3xFLAG™発現ベクターは、2つの高機能システムを1つの製品に統合したSnapFast™発現ベクターに変更されました。SnapFast™システムに関する追加情報には、背景情報、一般的に使われるベクター、選択チャート、SnapFast™に関するよくある質問が含まれます。 FLAG®および3xFLAG™ベクターは、T7、Tac、またはCMVプロモーターを使用する、細菌および哺乳類の発現システムで利用できます。ベクターには、精製されたタンパク質からタグを除去するためのエンテロキナーゼ切断部位があります。一部のベクターは、c-mycとともに、二重標識にも対応しています。ベクター仕様の概要については、以下のチャートを参照してください。ベクターマップなどの詳細については、製品ページのリンクをクリックしてください。 | | | | | | | | | | |-------------------------------------------------------------------|---------|---------|----------|--------------|---------|----------|---------|------------| | 製品番号 | FLAG®タグ | | OmpA分泌タグ | アンピシリン選択マーカー | EKT切断タグ | | | プロモーター | | | __N末端__ | __C末端__ | | | | __誘導物質__ | __TAC__ | __T7 Lac__ | | [OGS632](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS632) | ✓ | | | ✓ | ✓ | ✓ | ✓ | | | [OGS633](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS633) | ✓ | | ✓ | ✓ | ✓ | ✓ | ✓ | | | OGS634 | | ✓ | | ✓ | ✓ | ✓ | ✓ | | | [OGS635](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS635) | | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ | | | [OGS636](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS636) | ✓ | | | ✓ | ✓ | ✓ | | ✓ | 表1.FLAG®および3xFLAG™タグによる細菌ベクターオプション | | | | | | | | | | | | | |-------------------------------------------------------------------|---------|-----|-----|------------|---------|---------|---------|------------------|---|-------------------|--------| | 製品番号 | FLAG®タグ | | | 3X FLAG™タグ | c-Mycタグ | EKT切断タグ | PPT分泌タグ | アンピシリン選択マーカー（細菌） | | ネオマイシン選択マーカー（哺乳類） | プロモーター | | | N末端 | C末端 | N末端 | C末端 | C末端 | | | | | 構成型 | CMV | | [OGS618](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS618) | ✓ | | | | | ✓ | | ✓ | | ✓ | ✓ | | [OGS619](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS619) | | ✓ | | | | ✓ | | ✓ | | ✓ | ✓ | | [OGS620](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS620) | | | ✓ | | | ✓ | | ✓ | | ✓ | ✓ | | [OGS621](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS621) | | | ✓ | | | ✓ | ✓ | ✓ | | ✓ | ✓ | | [OGS622](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS622) | | | ✓ | | ✓ | ✓ | ✓ | ✓ | | ✓ | ✓ | | [OGS623](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS623) | | | ✓ | | ✓ | ✓ | | ✓ | | ✓ | ✓ | | [OGS624](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS624) | ✓ | | | | | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ | | [OGS625](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS625) | ✓ | | | | | ✓ | | ✓ | ✓ | ✓ | ✓ | | [OGS626](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS626) | | | ✓ | | | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ | | [OGS627](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS627) | | | ✓ | | | ✓ | | ✓ | ✓ | ✓ | ✓ | | [OGS628](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS628) | | | | ✓ | | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ | | [OGS629](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS629) | | | | ✓ | | ✓ | | ✓ | ✓ | ✓ | ✓ | | [OGS630](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS630) | | | ✓ | | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ | | [OGS631](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS631) | | | ✓ | | ✓ | ✓ | | ✓ | ✓ | ✓ | ✓ | 表2.FLAG®および3xFLAG™タグによる哺乳類ベクターオプション [](https://www.sigmaaldrich.com) ## FLAG®および3xFLAG™：タンパク質の検出と精製用に確立されたシステム 哺乳動物細胞における組換えタンパク質の低発現レベルは、エピトープタグの検出を困難または不可能にすることの多い一般的な問題です。ANTI-FLAG® M2 Abおよび抗マウスHRPを使用した3xFLAG™の検出感度は、一般的に使用されるタグを20〜200倍上回ります。選択の幅が広いp3xFLAG™ベクターでは、発現の促進に強力なCMVプロモーターが利用されています。  ウェスタンブロッティングを用い、精製された3xFLAG™細菌性アルカリホスファターゼとFLAG®細菌性アルカリホスファターゼをニトロセルロースメンブレンに転写しました。検出は、ANTI-FLAG® M2モノクローナル抗体（一次抗体）、抗マウス-HRP二次抗体、およびECL™化学発光基質を使用して行いました。  GSTのC末端融合体と上記のタグをウェスタンブロッティングで調製し、分析しました。適切なタグ抗体、抗マウスHRPおよびECLを使用して、すべてのタンパク質が検出されました。 [](https://www.sigmaaldrich.com) ## FLAG®および3xFLAG™エピトープに対する高特異性抗体 FLAG®および3xFLAG™配列には、それぞれ異なる認識性と結合性を持つ、非常に特異的なANTI-FLAG®モノクローナル（M1、M2、M5）およびポリクローナル抗体とコンジュゲートの結合部位が含まれています。ANTI-FLAG®抗体は、大半の哺乳類および細菌の細胞溶解液において交差反応性をほとんどまたはまったく示しません。ワンステップ精製フォーマットには、アフィニティゲルと96ウェルプレートが含まれます。 ## [](https://www.sigmaaldrich.com)SnapFast™ FLAG®および3xFLAG™ベクター従来製品比較表 注記：過去にFLAG®ベクターを使用したことがある場合は、このチャートを使用して、それに最も近いSnapFast™ FLAG®および3xFLAG™ベクターを選択できます。 | | | |-------------------------------------------------------------------|----------------------------------------------------------------------------------------------------------------------------------| | SnapFast™ FLAG®または3xFLAG™ベクター製品番号 | 従来のFLAG®ベクター製品番号 | | [OGS618](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS618) | [E6783](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E6783) | | [OGS619](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS619) | [E6908](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E6908) | | [OGS620](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS620) | [E7033](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E7033) | | [OGS621](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS621) | [E7158](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E7158) | | [OGS622](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS622) | [E7283](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E7283) | | [OGS623](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS623) | [E9283](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E9283) | | [OGS626](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS626) | [E7658](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E7658) ・[E9783](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E9783) | | [OGS627](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS627) | [E7408](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E7408) ・[E7533](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E7533) | | [OGS628](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS628) | [E7783](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E7783) ・[E9908](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E9908) | | [OGS629](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS629) | [E7908](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E7908) | | [OGS630](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS630) | [E9158](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E9158) ・[E9408](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E9408) | | [OGS631](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS631) | [E9283](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E9283) | | [OGS632](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS632) | E8083 | | [OGS633](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS633) | [E8158](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E8158) | | OGS634 | [E8408](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E8408) | | [OGS635](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS635) | [E8283](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/E8283) | | [OGS636](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/OGS636) | [P1118](https://www.sigmaaldrich.com/JP/ja/product/SIGMA/P1118) | __関連資料__ - [細胞へのトランスフェクション入門](https://www.sigmaaldrich.com/JP/ja/technical-documents/technical-article/cell-culture-and-cell-culture-analysis/transfection-and-gene-editing/transfection-reagents) - [組換えタンパク質の超高感度検出を可能にする3xFLAG®システム発現ベクター](https://www.sigmaaldrich.com/JP/ja/technical-documents/technical-article/genomics/cloning-and-expression/3x-flag) - [コロニー選択のための青／白スクリーニングとそのプロトコル](https://www.sigmaaldrich.com/JP/ja/technical-documents/technical-article/genomics/cloning-and-expression/blue-white-screening) - 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