Invitrogen™ Bac-to-Bac™ N-His TOPO™ Expression System

Bac-to-Bac™ N-His TOPO™ Expression System

Supplier:  Invitrogen™ A11101

Catalog No. A11101

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. Bac-to-Bac™ relies on generation of recombinant baculovirus by 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. Approximately 100 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 line 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

• Type: Expression System
• Protein Tag: His Tag (6x)
• Content And Storage: Bac-to-Bac N-His TOPO Expression Kit:
  • Vector kit for 20 TOPO cloning reactions, store at -20°C
     –pFastBac/NT-TOPO vector containing N-terminal His-tag and TEV cleavage site
     –pFastBac 1-Gus control plasmid (control expression vector)
     –Other kit reagents: 10X PCR, dNTP mix, Salt solution, sterile water, control PCR template, polyhedrin forward primer, SV40 pA reverse primer
  • One Shot Mach1-T1R Chemically Competent E. coli for 20 reactions (20 x 50 µL), store at -80°C
  • MAX Efficiency DH10Bac Competent E. coli for 20 reactions (4 kits with 5 x 0.1 mL each), store at -80°C
  • ExpiFectamine Sf Transfection Reagent (1 mL), store at 4°C, do not freeze
  
  
• Format: Liquid
• Expression System: Baculovirus
• Promoter: Polyhedrin

• Cloning Method: Blunt TOPO™
• Quantity: 20 Reactions
• Antibiotic Resistance Bacterial: Ampicillin (AmpR), Gentamicin (GmR)
• Expression Mechanism: Cell-Based Expression
• Product Line: Bac-to-Bac™, TOPO™

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