规格
Cleavage:	TEV Protease Recognition Site
Cloning Method:	Gateway®
Product Line:	BaculoDirect™, Gateway®
Promoter:	Polyhedrin
Vector:	BaculoDirect Vectors
Protein Tag:	His Tag (6x), V5 Epitope Tag

储存

The BaculoDirect expression kits include five 0.3 µg vials of BaculoDirect Linear DNA, 125 µL of Cellfectin transfection reagent, frozen Sf9 cells, Gateway LR Clonase enzyme mix, GIBCO Unsupplemented Grace's Insect Medium, and ganciclovir.

Store the BaculoDirect Linear DNA, Cellfectin reagent, and Grace's Medium at +4°C. Store LR Clonase mix at -80°C. Store Sf9 cells in liquid nitrogen. All components are guaranteed stable for 6 months if stored properly.

描述

The BaculoDirect™ Baculovirus Expression System is a powerful and versatile eukaryotic system for high-level protein expression in insect cells. The combination of Gateway™ Technology with baculovirus expression makes the BaculoDirect System the fastest and easiest method for production of recombinant baculovirus.

How it works
BaculoDirect™ Linear DNA (the baculovirus genome) is engineered to include attR sites for quick and efficient recombination with a Gateway entry clone. The gene of interest is recombined from the entry clone into the BaculoDirect Linear DNA using a simple, one-hour LR reaction (Figure 1). The resulting reaction mix contains the recombinant baculovirus carrying the gene of interest and is used to transfect insect cells. The need for transforming bacteria and isolating a large bacmid or co-transfecting a transfer vector and linear baculovirus DNA into insect cells is eliminated. As a result, the hands-on time is greatly reduced. Purified baculovirus can be isolated in less than one week.

Gateway linear DNA
BaculoDirect Linear DNA is designed for rapid cloning with a Gateway entry clone and subsequent expression in Sf9 or Sf21 insect cells. The Linear DNA features:

• Strong polyhedrin promoter for high-level expression
• R-sites for efficient recombination with any attL-flanked Gateway entry vector
• TK gene for negative selection using ganciclovir
• C-terminal V5-His tag (BaculoDirect™ C-Term Expression Kit) or N-terminal V5-His tag (BaculoDirect™ N-Term Expression Kit) for detection with anti-V5 antibody and purification with nickel-chelating resin
• TEV protease cleavage site for removal of the V5-His tag following purification (BaculoDirect N-Term Expression Kit)
• LacZ gene, to ensure a pure baculovirus stock is generated, which is replaced by your gene of interest after LR recombinant reaction

Additional materials required, available separately: Gateway entry clone.

A selection guide for choosing the most appropriate Gateway entry vector for your application can be found at: www.thermofisher.com/Gateway.

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