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

Bac-to-Bac™ N-His TOPO™ Cloning Kit

Supplier:  Invitrogen™ A11099

Catalog No. A11099

Description

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

• Antibiotic Resistance Bacterial: Ampicillin (AmpR), Gentamicin (GmR)
• Cell Type: Competent Cell
• Content And Storage: 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
  
• Expression System: Baculovirus
• No. of Reactions: 20 Reactions
• Product Type: Cloning Kit
• Protein Tag: His Tag (6x)
• Selection Agent (Eukaryotic): Ampicillin, Gentamicin

• Cell Line: High Five™, Sf21, Sf9
• Cloning Method: Blunt TOPO
• Expression Mechanism: Insect Cell Expression
• For Use With (Application): Protein Expression, Viral Vaccine Production
• Product Line: Bac-to-Bac™, TOPO™
• Promoter: Polyhedrin
• Quantity: 20 Reactions
• Vector: pFastBac/NT-TOPO

For Research Use Only. Not for use in diagnostic procedures.