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Invitrogen™ Bac-to-Bac™ N-His TOPO™ Cloning Kit

Bac-to-Bac™ N-His TOPO™ Cloning Kit

Supplier:  Invitrogen™ A11099


Catalog No. A11099

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Bac-to-Bac™ Baculovirus Expression is an efficient method for producing baculovirus for high-level protein expression in insect cells. It relies on generation of recombinant virus by site-specific transposition in E. coli rather than homologous recombination in insect cells. New: pFastBacTOPO™ vectors!

Reliable and fast protein expression
The Bac-to-Bac™ Baculovirus Expression System is faster and easier than traditional baculovirus expression, and it maintains high levels of protein expression. It is based on site-specific transposition in E. coli rather than homologous recombination in insect cells. The Bac-to-Bac™ Baculvirus Expression System is highly regarded in academic Literature. At least 80 citations every year since 2000!

Traditional baculovirus systems require purification and amplification of an initial low-titer viral supernatant. This requires a time-consuming plaque assay. Bac-to-Bac™'s pFastBac™ vector, however, recombines with the parent bacmid in DH10Bac™ E. coli competent cells to form an expression bacmid. Transfect the bacmid into insect cells for fast production of a high titer of pure recombinant baculovirus particles in the very first transfection. You'll save weeks of precious time. Collect a pure P2 baculovirus stock on Day 10 without the necessity of tedious plaques assay (See Figure 1)

High expression, easy screening
pFastBac™ uses the strong polyhedrin promoter to generate high levels of expression in a variety of insect cell lines such as Sf9, Sf21, and High Five™ cells.

New additions:
Bac-to-Bac™'s pFastBac vectors are now available as pFastBacTOPO™ vectors too. pFastBacTOPO™ vectors are Blunt™ TOPO™ vectors and are supplied with Mach1™-T1R E. coli for easy cloning and pFastBac⁄GOI propagation. The new vectors enable you to:

  • Amplify your gene of interest (GOI) with a PCR enzyme of highest fidelity such as Accuprime™ Pfx SuperMix.

  • TOPO™ clone the blunt-end PCR product into the new vector in only 5 minutes!

  • Visualize colonies 8 hours after plating on ampicillin selective plates because Mach1™-T1R E. coli cells have a faster doubling time compared to other standard cloning strains.

  • With the Bac-to-Bac™ N-His TOPO™ cloning or expression kit, you can produce a N-terminal His-fusion protein with a TEV cleavage site to purify with nickel-chelating resin (Invitrogen’s ProBond™ Purification System) and generate a native protein with the aid of Invitrogen’s popular AcTEV™ Protease.

    The new vectors combine the TOPO™ cloning technology with the highly regarded Bac-to-Bac™ baculovirus expression features to enable easy cloning and high-level protein expression.
    The pFastBacTOPO™ vectors are offered as cloning and expression kits.
  • Specifications


    Ampicillin (AmpR), Gentamicin (GmR)
    Competent Cell
    Kit content for Bac-to-Bac™ N-His TOPO™ cloning kit (For details see the manual)
    • Vector kit for 20 topo cloning reactions:
    o pFastBac⁄NT-TOPO™ Vector containing the N-terminal His-Tag and TEV cleavage site.
    o pFastBac⁄NT-Gus Control Plasmid (Control expression vector)
    o Other reagents supplied: 10x PCR, dNTP Mix, Salt Solution, Sterile Water, Control PCR template, Polyhedrin Forward Primer, SV40 pA Reverse Primer
    • Kit containing competent cells (20 reactions):
    o One Shot™ Mach1-T1R Chemically Competent E. coli
    Vector kit: store at -20°C
    One Shot™ Mach1-T1R Chemically Competent E. coli: store at -80°C
    20 Reactions
    Cloning Kit
    His Tag (6x)
    Ampicillin, Gentamicin
    High Five™, Sf21, Sf9
    Blunt TOPO
    Insect Cell Expression
    Protein Expression, Viral Vaccine Production
    Bac-to-Bac™, TOPO™
    20 Reactions


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