ViraPower™ HiPerform™ T-REx™ Gateway® Expression System

Catalog number: A11141

Invitrogen™  Related applications: Protein Expression

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Description

The ViraPower™ HiPerform™ T-REx™ Gateway® Expression System includes all the components needed to generate lentivirus, including vector kit, 293FT cell line, and the support kit. This kit combines Invitrogen’s ViraPower™ HiPerform™ Lentiviral, T-REx™ and Gateway® technologies to facilitate easy recombination-based cloning and lentiviral-based, regulated (Tetracycline-inducible), high-level expression of a target gene in dividing and non-dividing mammalian cells. The pLenti6.3⁄ TO⁄ V5-DEST vector is equipped with two key genetic elements, making it a HiPerform™ vector: the Woodchuck Posttranscriptional Regulatory Element (WPRE) and the central Polypurine Tract (cPPT) sequence from the HIV-1 integrase gene to produce at least 4-fold increase in protein expression compared to vectors lacking these elements.

Advantages
• Generates replication-incompetent lentivirus for transducing dividing and non-dividing mammalian cells
• Easy recombination-based cloning using Gateway® technology
• Stable, long-term, tetracycline-regulated expression
• Enhanced protein expression, up to 4-fold or greater, compared to traditional lentiviral expression systems

Key Features
• WPRE (Woodchuck Posttranscriptional Regulatory Element) from the woodchuck hepatitis virus, increases transgene expression and cPPT (central Polypurine Tract) from the HIV-1 integrase gene, increases the copy number of lentivirus integrating into the host genome, thus increasing viral titer. WPRE and cPPT together produce at least a four-fold increase in protein expression in most cell types, compared to other vectors that do not contain these elements.
• Hybrid promoter consisting of the human cytomegalovirus (CMV) promoter and two tetracycline operator 2 (TetO2) sites for high-level, regulated expression of the target gene
• Blasticidin selection marker for stable selection under control of SV40 promoter

Kit Includes
• ViraPower™ HiPerform™ T-Rex™ Gateway® Vector Kit (Cat # A11144)
• ViraPower™ Bsd Lentiviral Support Kit (Cat # K497000)
• 293FT Cell Line (Cat # R70007)
• Gateway® LR Clonase® II Plus Enzyme Mix (Cat # 12538120)
• One Shot® Stbl3™ Chemically Competent E. coli (Cat # C737303)
• Geneticin® (Cat # 10131035)
• pENTR™ Gus positive control plasmid

For research use only. Not intended for any therapeutic or diagnostic use.
For Research Use Only. Not for use in diagnostic procedures.

Specifications

Promoter: CMV⁄TO
Key Function: Viral Expression
Protein Tag or Fusion: V5 Epitope Tag
Delivery Method: Lentiviral
Cloning Method: Gateway®
Selection Agent (Eukaryotic): Blasticidin
Vector Type: pLenti
Antibiotic Resistance (Bacterial): Ampicillin (AmpR), Blasticidin (BsdR)
Cleavage: No Cleavage Site
Constitutive or Inducible System: Inducible
Gateway Enzyme: LR Clonase® II, LR Clonase® II Plus
Inducing Agent: Doxycycline, Tetracycline
PCR Enzyme: Proof-Reading Polymerase
Product Size: 20 reactions
Protein Tag Position (to your gene): C-terminal
Replication Origin: pUC Origin
Selection Marker (Eukaryotic): BsdR
Selection Marker Promoter: PGK Promoter
Special Elements: WPRE, cPPT

Contents & storage

Contents

The ViraPower™ HiPerform™ T-Rex™ Gateway® Expression System includes the following components:
• ViraPower™ HiPerform™ T-Rex™ Gateway® Vector Kit (Cat # A11144) which includes:
  • 6 ug of pLenti6.3⁄TO⁄V5-DEST vector (40 µL of vector at 150 ng⁄µL in TE Buffer, pH 8.0)
  • 20 ug of pLenti3.3⁄TR vector (40 µL of vector at 0.5 µg⁄µL in TE Buffer, pH 8.0)
  • 10 ug of pLenti6.3⁄TO⁄V5-GW⁄lacZ plasmid (20 µL of plasmid at 0.5 µg⁄µL in TE Buffer, pH 8.0)
  • 1 ml of Tetracycline (10 mg⁄mL in water)

  • • ViraPower™ Bsd Lentiviral Support Kit (Cat # K497000) which includes:
  • 195 ug of ViraPower™ Lentiviral Packaging Mix (Contains 195 uL of a mixture of pLP1, pLP2, and pLP⁄VSVG plasmids, at 1 ug⁄uL in TE Buffer, pH 8.0)
  • 0.75 ml of Lipofectamine® 2000 (Cat # 11668027)
  • 50 mg of Blasticidin powder (Cat # R21001)

  • • 293FT Cell Line (Cat # R70007): 1 vial containing 3 X 106 frozen cells in 1 mL of Freezing Medium.
    • Gateway® LR Clonase® II Plus Enzyme Mix (Cat # 12538120): 40 uL of Gateway® LR Clonase® II Plus Enzyme Mix and 40 uL of Proteinase K Solution at a concentration of 2 ug⁄mL.
    • One Shot® Stbl3™ Chemically Competent E. coli (Cat # C737303): 21 X 50 uL of Stbl3™ Cells, 6 mL of S.O.C. Medium and 50 uL of pUC19 control DNA (10 pg⁄uL).
    • Geneticin® (Cat # 10131035): 20 mL of Geneticin® solution (50 mg⁄mL).
    • pENTR™ Gus positive control plasmid

    Storage

    • Vectors: -20°C
    • ViraPower™ Packaging Mix: -20°C
    • LR Clonase® II Plus Enzyme Mix: -20°C for storage up to 6 months, -80°C for long term storage
    • Tetracycline: -20°C (protected from light)
    • Blasticidin: -20°C
    • Geneticin: 4°C or -20°C
    • Lipofectamine® 2000: 4°C (do not freeze)
    • One Shot® Stbl3™ Chemically Competent E.coli: -80°C
    • 293 FT Cells: Liquid Nitrogen
    • pENTR™ Gus positive Control: -20°C

    Documents

    Manuals & protocols