Promoterless NanoLuc® Genetic Reporter Vectors with Hygromycin Selection
Clone a Putative Promoter for Expression in Mammalian Cells and Select Stable Transfectants
- Offers two engineered genes for your reporter assay: Nluc and NlucP
- NanoLuc® enzyme and its substrate are about 100-fold brighter than other luciferases, producing a high intensity, glow-type luminescence
- Use for transient transfection or select for stable expression with hygromycin
Catalog Number:
Choose a luciferase gene
Size
Catalog Number: N1061
Catalog Number: N1071
レポーター遺伝子としてのNanoLuc® ルシフェラーゼは複数の形態を用意しており、実験目的に合うように構成されています。標準型のNlucは最大の発光出力と感度を備えています。NanoLuc®-PEST（NlucP）は転写活性の変化とタンパク質発現がより密接に対応しており、シグナル/バックグラウンド比が向上します。分泌型レポーターが必要な場合にはN末端に分泌シグナルが融合されたNanoLuc® ルシフェラーゼ（secNluc）も用意しています。発光値はNano-Glo® Luciferase Assay Reagent で測定した場合、NanoLuc®タンパク質量と1,000,000倍以上の濃度幅で比例し、シグナルの半減期は2時間以上です。
NanoLuc® ルシフェラーゼは優れたレポータータンパク質として数多くの物理的特性を備えています
• 非常に小さい、単量体酵素（171 アミノ酸；513bp）
• 高い熱安定性（Tm = 60℃）
• 広範なpH幅で活性を保持（pH 6 - 8）
• 翻訳後修飾、ジスルフィド結合がない
• 細胞内で均一に分布
• 蛍光スペクトルが生物発光共鳴エネルギー転移（BRET；λmax = 465nM）に最適
NanoLuc® ルシフェラーゼは転写制御のレポーターアッセイで使用するためにデザインされた様々な構成のプラスミド（標準型 Nluc, PEST 不安定型 NlucP, 分泌型 secNluc）がご利用いただけます。
各pNLベクターの用途：
• pNL1：既知または推定されるプロモーター領域のクローニング
• pNL2：既知または推定されるプロモーター領域のクローニングおよびハイグロマイシン選択による安定細胞株の構築
• pNL3：基本プロモーターを含まない結合サイトや応答エレメントのクローニング（pNL3.2.NF-κB-RE vector内に存在するような）
• 標準型および分泌型 Nluc 用のコントロールプラスミドも用意
pNLベクターシリーズはpGL4の基本骨格を用いているため既存のプラスミドからのサブクローニングが容易です。この骨格は数多くの転写因子結合サイトやその他の潜在的な調節エレメンを除去することにより変則的な結果を低減するようデザインされています。Nluc 遺伝子はコドンの最適化や多くの潜在的な調節エレメント、その他の不要な特徴が除かれています（例：一般的な制限酵素サイトなど）。
- 転写調節
- ウイルス/細胞間相互作用
- 化合物スクリーニング
- 翻訳後修飾
- GPCRシグナリング
- 細胞内シグナリング
- プロモーター解析
【ライセンス】NanoLuc
【Ｑ＆Ａ】Reporter_1.
About NanoLuc® Luciferase and the pNL Vectors
For use as a genetic reporter, multiple forms of NanoLuc® luciferase have been configured to meet differing experimental objectives. Unfused Nluc offers maximal light output and sensitivity, and NanoLuc®-PEST (NlucP) closely couples protein expression to changes in transcriptional activity and increased signal-to background ratios. When a secreted reporter is preferred, there is NanoLuc® luciferase fused to an N-terminal secretion signal (secNluc). Luminescence is linearly proportional to the amount of NanoLuc® protein over a 1,000,000-fold concentration range, with a signal half-life ≥2 hours when detected with Nano-Glo® Luciferase Assay Reagent.
NanoLuc® luciferase possesses a number of physical properties that make it an excellent reporter protein:
- very small, monomeric enzyme (171 amino acids; 513bp)
- high thermal stability (Tm = 60°C)
- active over a broad pH range (pH 6–8)
- no post-translational modifications or disulfide bonds
- uniform distribution in cells
- emission spectrum well suited for bioluminescence resonance energy transfer (BRET; λmax = 465nM).
NanoLuc® luciferase is made available in a variety of plasmids designed for use in reporter gene assays of transcriptional control and with each of the NanoLuc® forms (unfused Nluc, PEST destabilized NlucP, and secreted secNluc). The different pNL variations are designed for the following:
- pNL1: cloning of a known or putative promoter region
- pNL2: cloning of a known or putative promoter region and establishment of a stable cell line through Hygromycin selection
- pNL3: cloning of a binding site or response element not in need of a basic promoter (such as are present in the pNL3.2.NF-κB-RE vector)
- Control plasmids for the unfused, PEST-destabilized and secreted Nluc forms also are available.
The pNL vector series uses a pGL4-based backbone for easy sequence transfer from existing plasmids. This backbone design also reduces anomalous results by removing many transcription factor binding sites and other potential regulatory elements. The Nluc gene variations are codon optimized and have had many potential regulatory elements or other undesirable features removed (such as common restriction enzyme sites).
NanoLuc® Technology performs similarly to firefly luciferase but gives higher light output.
pNL2.1[Nluc/Hygro] Vector GenBank® Accession Number JQ513372 and vector sequence text file.
pNL2.2[NlucP/Hygro] Vector GenBank® Accession Number JQ513373 and vector sequence text file.
A comparison of light output and response dynamics between NanoLuc® and firefly luciferase constructs.
Protocols
No protocols available
Specifications
Catalog Number:
選択製品の構成品内容
| Item | Part # | Size |
|---|---|---|
| pNL2.1[Nluc/Hygro] Vector | N106A | 1 × 20μg |
BY USE OF THIS PRODUCT, RESEARCHER AGREES TO BE BOUND BY THE TERMS OF THIS LIMITED USE LABEL LICENSE. If researcher is not willing to accept the conditions of this limited use statement, and the product is unused, Promega will accept return of the unused product and provide researcher with a full refund.
Researchers may use this product for research use only; no transfer or commercial use of this product is allowed. Commercial use means any and all uses of this product or derivatives by a party in exchange for consideration, including, but not limited to (1) use in further product manufacture; (2) use in provision of services, information or data; and (3) resale of the product or its derivatives, whether or not such product or derivatives are resold for use in research. Researchers shall have no right to modify or otherwise create variations of the nucleotide sequence of the luciferase gene except that researcher may (1) create fused gene sequences provided that the coding sequence of the resulting luciferase gene has no more than four deoxynucleotides missing at the affected terminus compared to the intact luciferase gene sequence; and (2) insert and remove nucleic acid sequences in splicing research predicated on the inactivation or reconstitution of the luminescence of the encoded luciferase. No other use of this product or derivatives is authorized without the prior express written consent of Promega.
In addition, researchers must:
(1a) use Nano-Glo®-branded luminescent assay reagents (LARs) for all determinations of luminescence activity of this product and its derivatives; or
(1b) contact Promega to obtain a license for use of the product and its derivatives with LARs not manufactured by Promega.
For uses of Nano-Glo®-branded LARs intended for energy transfer (such as bioluminescence resonance energy transfer) to acceptors other than a genetically encoded autofluorescent protein, researchers must:
(2a) use NanoBRET®-branded energy acceptors (e.g., BRET-optimized HaloTag® ligands) for all determinations of energy transfer activity by this product and its derivatives; or
(2b) contact Promega to obtain a license for use of the product and its derivatives for energy transfer assays to energy acceptors not manufactured by Promega.
Researchers may transfer derivatives to others for research use provided that at the time of transfer a copy of this label license is given to the recipients and recipients agree to be bound by the terms of this label license. With respect to any uses outside this label license, including any diagnostic, therapeutic, prophylactic or commercial uses, please contact Promega for supply and licensing information. PROMEGA MAKES NO REPRESENTATIONS OR WARRANTIES OF ANY KIND, EITHER EXPRESSED OR IMPLIED, INCLUDING FOR MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE, WITH REGARD TO THE PRODUCT. The terms of this agreement shall be governed under the laws of the State of Wisconsin, USA.
U.S. Pat. Nos. 8,557,970, 8,669,103, 9,777,311, 9,840,730, 9,951,373, 10,233,485, 10,633,690, 10,774,364, 10,844,422, 11,365,436, 11,661,623, 11,667,950; European Pat. Nos. 2456864, 2635595, 2990478, 3181687, 3409764; Japanese Pat. Nos. 6038649, 6155424, 6227615, 6374420, 6539689; and other patents and patents pending.
選択製品の構成品内容
| Item | Part # | Size |
|---|---|---|
| pNL2.2[NlucP/Hygro] Vector | N107A | 1 × 20μg |
BY USE OF THIS PRODUCT, RESEARCHER AGREES TO BE BOUND BY THE TERMS OF THIS LIMITED USE LABEL LICENSE. If researcher is not willing to accept the conditions of this limited use statement, and the product is unused, Promega will accept return of the unused product and provide researcher with a full refund.
Researchers may use this product for research use only; no transfer or commercial use of this product is allowed. Commercial use means any and all uses of this product or derivatives by a party in exchange for consideration, including, but not limited to (1) use in further product manufacture; (2) use in provision of services, information or data; and (3) resale of the product or its derivatives, whether or not such product or derivatives are resold for use in research. Researchers shall have no right to modify or otherwise create variations of the nucleotide sequence of the luciferase gene except that researcher may (1) create fused gene sequences provided that the coding sequence of the resulting luciferase gene has no more than four deoxynucleotides missing at the affected terminus compared to the intact luciferase gene sequence; and (2) insert and remove nucleic acid sequences in splicing research predicated on the inactivation or reconstitution of the luminescence of the encoded luciferase. No other use of this product or derivatives is authorized without the prior express written consent of Promega.
In addition, researchers must:
(1a) use Nano-Glo®-branded luminescent assay reagents (LARs) for all determinations of luminescence activity of this product and its derivatives; or
(1b) contact Promega to obtain a license for use of the product and its derivatives with LARs not manufactured by Promega.
For uses of Nano-Glo®-branded LARs intended for energy transfer (such as bioluminescence resonance energy transfer) to acceptors other than a genetically encoded autofluorescent protein, researchers must:
(2a) use NanoBRET®-branded energy acceptors (e.g., BRET-optimized HaloTag® ligands) for all determinations of energy transfer activity by this product and its derivatives; or
(2b) contact Promega to obtain a license for use of the product and its derivatives for energy transfer assays to energy acceptors not manufactured by Promega.
Researchers may transfer derivatives to others for research use provided that at the time of transfer a copy of this label license is given to the recipients and recipients agree to be bound by the terms of this label license. With respect to any uses outside this label license, including any diagnostic, therapeutic, prophylactic or commercial uses, please contact Promega for supply and licensing information. PROMEGA MAKES NO REPRESENTATIONS OR WARRANTIES OF ANY KIND, EITHER EXPRESSED OR IMPLIED, INCLUDING FOR MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE, WITH REGARD TO THE PRODUCT. The terms of this agreement shall be governed under the laws of the State of Wisconsin, USA.
U.S. Pat. Nos. 8,557,970, 8,669,103, 9,777,311, 9,840,730, 9,951,373, 10,233,485, 10,633,690, 10,774,364, 10,844,422, 11,365,436, 11,661,623, 11,667,950; European Pat. Nos. 2456864, 2635595, 2990478, 3181687, 3409764; Japanese Pat. Nos. 6038649, 6155424, 6227615, 6374420, 6539689; and other patents and patents pending.
Resources
No related resources available
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