Invitrogen Bac-to-Bac HBM TOPO Secreted Expression System
Bac-to-Bac HBM TOPO Secreted Expression System
Manufacturer: Invitrogen A11339
DescriptionBac-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 (DH10Bac) rather than homologous recombination in insect cells. The new pFastBac HBM TOPO vector enables secreted protein expression because it has the honeybee melittin (HBM) secretion signal. The new vector should be strongly considered for the expression of glycoproteins. Glycoproteins cannot be glycosylated in the absence of any secretion signal. In contrast to glycoproteins secreted from mammalian cells, glycoproteins secreted from baculovirus can be easily de-glycosylated in vitro. This is a very important feature in order to crystallize proteins!
The pFastBac HBM TOPO vector has also a C-terminal His-Tag with a TEV cleavage signal to enable easy purification of Histidine fusion proteins on nickel-chelating resins (ProBond Purification System) and native proteins with the aid of AcTev protease.
The vector uses the strong polyhedrin promoter to generate high levels of expression in a variety of insect cell lines such as Sf9, Mimic Sf9, Sf21, and High Five cells in Sf-900 II & Sf-900 III media from Gibco.
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 vectors, 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.
|Bac-to-Bac™ HBM TOPO™ Secreted Expression System, 20rxns, Liquid, TEV Protease Recognition Site Cleavage, Blunt TOPO™ Cloning Method, Baculovirus, Insect Expression, Polyhedrin Promoter, His Tag Fusion, Cell-Based Expression Mechanism, Used for Protein Expression|
|Protein biology, protein expression, insect expression, high-level protein production in insect cells, secreted protein expression in insect cells, baculovirus production and titering|
|–30 to –10°C|
|Bac-to-Bac™ HBM TOPO™ Secreted Expression System|
|Vector kit for 20 topo cloning reactions, pFastBac/HBM-TOPO™ Vector containing the C-terminal TEV cleavage sit and His-Tag, pFastBac/CT-Gus control plasmid (Control expression vector), 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), One Shot™ Mach1-T1R chemically competent E. coli, 4 kits with 5 x 0.1mL each of MAX Efficiency™ DH10Bac™ competent E. coli for 20 reactions, 1mL Cellfectin™ II reagent, Vector kit, One shot™ Mach1-T1R chemically competent E. coli, MAX Efficiency™ DH10Bac™ competent E. coli, Cellfectin™ II Reagent: store at 4°C|
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