ViraPower™ HiPerform™ Promoterless Gateway® Vector Kit

Catalog number: A11146

Invitrogen™  Related applications: Protein Expression

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Description

The ViraPower™ HiPerform™ Promoterless Gateway® Vector Kit contains the MultiSite Gateway®-adapted ViraPower™ HiPerform™ promoterless lentiviral expression vector, pLenti6.4⁄R4R2⁄V5-DEST™ for easy recombination-based cloning and lentiviral-based high-level expression of a target gene from any promoter of choice in dividing and non-dividing mammalian cells. The pLenti6.4⁄R4R2⁄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, simultaneous, recombination-based cloning of multiple DNA fragments in a defined order and orientation using MultiSite Gateway® technology
• Expression of the target gene under the control of a promoter of choice
• Stable, long-term expression
• Enhanced protein expression, up to 4-fold or greater, compared to traditional lentiviral expression systems

Key Features
• WPRE from the woodchuck hepatitis virus, increases transgene expression and cPPT 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.
• Promoterless vector to express the target gene under the control of a promoter of choice
• Blasticidin selection marker for stable selection under control of PGK promoter for long-term, persistent expression

Kit includes
• pLenti6.4⁄R4R2⁄V5-DEST™ Kit
• pENTR™ 5’ TOPO® TA Cloning Kit (Cat # K59120)
• One Shot® Stbl3™ Chemically Competent E. coli (Cat # C737303)
• 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: None (Promoterless), CMV, EF-1α
Key Function: Viral Expression, Clone your own Promoter
Protein Tag or Fusion: V5 Epitope Tag
Delivery Method: Lentiviral
Cloning Method: Gateway®
Selection Agent (Eukaryotic): Blasticidin
Vector Type: pLenti
Cleavage: No Cleavage Site
Comments: Use the provided EF1&alpha or CMV promoter, or clone your own promoter of interest.
PCR Enzyme: Proof-Reading Polymerase
Product Size: 20 reactions
Gateway Enzyme: LR Clonase® II, LR Clonase® II Plus
Special Elements: WPRE, cPPT
Replication Origin: pUC Origin
Selection Marker Promoter: PGK Promoter
Selection Marker (Eukaryotic): BsdR
Constitutive or Inducible System: Constitutive
Antibiotic Resistance (Bacterial): Ampicillin (AmpR), Blasticidin (BsdR)
Protein Tag Position (to your gene): C-terminal

Contents & storage

Contents
The ViraPower™ HiPerform™ Promoterless Gateway® Vector Kit includes the following components:

• pLenti6.4⁄R4R2⁄V5-DEST™ Kit which includes:
  • 6 µg of pLenti6.4⁄R4R2⁄V5-DEST™ vector (40 µL of vector at 150 ng⁄µL in TE Buffer, pH 8.0)
  • 10 µg of pLenti6.4⁄CMV⁄V5-MSGW⁄lacZ plasmid (20 µL of vector at 0.5 µg⁄µL in TE Buffer, pH 8.0)
  • 10 µg of pENTR 5’⁄CMVp plasmid (20 µL of plasmid at 0.5 µg⁄µL in TE Buffer, pH 8.0)
  • 10 µg of pENTR 5’⁄EF1αp plasmid (20 µL of plasmid at 0.5 µg⁄µL in TE Buffer, pH 8.0)

  • • pENTR 5’ TOPO® TA Cloning Kit (Cat # K95120) which includes:
  • pENTR 5’ TOPO® Reagents
  • One Shot® TOP10 Chemically Competent E.coli (Cat # C404003)
  • One Shot® Stbl3™ Chemically Competent E. coli (Cat # C737303): 21 X 50 µL of Stbl3™ Cells, 6 mL of S.O.C. Medium and 50 µL of pUC19 control DNA (10 pg⁄µL)
  • pENTR™ Gus positive control plasmid

  • Storage
    • pLenti6.4⁄R4R2⁄V5-DEST™ Kit: -20°C
    • pENTR™ 5’ TOPO® Reagents: -20°C
    • One Shot® Stbl3™ Chemically Competent E.coli: -80°C
    • One Shot® TOP10 Chemically competent E.coli: -80°C
    • pENTR™ Gus positive Control: -20°C

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