Shop All Protein Expression Kits

Bac-to-Bac™ C-His TOPO™ Cloning Kit (Gibco™)

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 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® C-His TOPO® cloning or expression kit, you can produce a C-terminal His-fusion protein with a TEV cleavage site to purify with nickel-chelating resins (Invitrogen’s ProBond™ Purification System) and generate a native protein with the aid of Invitrogen’s popular AcTEV™ Protease. This is the Invitrogen’s only vector that contains a C-terminal TEV cleavage site!

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.

Champion™ pET104 BioEase™ Gateway™ Biotinylation System (Invitrogen™)

The Champion™ pET104-DEST BioEase™ Gateway® Expression System provides an easy, efficient method for expressing, purifying, and detecting biotinylated recombinant proteins. The Champion™ pET104-DEST vector (Figure 1) incorporates a 72 amino acid sequence from K. pneumoniae that directs in vivo biotinylation of a specific lysine residue. This tag is recognized and efficiently biotinylated by the enzyme Biotin Protein Ligase (BPL) encoded by the birA gene in E. coli(1). Proteins produced in the Champion™ pET104-DEST vector are expressed as fusion proteins to this sequence. Biotinylated fusion proteins can be purified on streptavidin agarose or detected on western blots using streptavidin-HRP (Figure 2) or streptavidin-AP conjugates.

In addition to the BioEase™ tag, the Champion™ pET104-DEST vector includes the following features:

attR sites for efficient recombination with any attL-flanked Gateway® entry vector
• The T7lac bacteriophage promoter for high-level expression of the recombinant fusion protein
• An enterokinase cleavage site for removal of the N-terminal BioEase™ fusion tag

MembraneMax™ HN Protein Expression Kit (Invitrogen™)

MembraneMax™ HN Protein Expression Kits delivers high yields of soluble (dispersed) membrane proteins using the MembraneMax™ reagent, which is an planar phospholipid membrane bilayer surrounded by a scaffold protein (also called a nanolipoprotein particle or NLP) that also offers convenient His-tag purification. Combining this technology with a scalable cell-free expression E. coli extract delivers microgram to milligram quantities of your membrane protein specifically for high throughput expression screening, expression of membrane proteins that would otherwise be toxic in cell-based systems and functional or activity assays. With high uniformity and consistent structure, MembraneMax reagent minimizes formation of membrane protein aggregates or clumping often caused by inconsistent size, shape and structure typical observed with microsomes or micelles. The MembraneMax™ Protein Expression kits contain all the necessary reagents for successful expression of your membrane protein.
Key Advantages of the MembraneMax™ Protein Expression Kits:
• Optimized cell-free expression delivers high yield of membrane protein from nanogram to milligram quantities
• MembraneMax™ reagent enables a monodispersed population of soluble membrane protein-NLP complexes
• Convenient His-tag allows for simple purification of native or un-tagged membrane proteins
• Expression format amenable to high-throughput protein synthesis for screening and expression of toxic proteins
• Production of a homogeneous population of your membrane protein
• Convenient kit format includes the necessary reagents for protein expression—simply add your gene of interest to get started

Bac-to-Bac™ N-His TOPO™ Cloning Kit (Gibco™)

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.

NativePure™ Lentiviral Expression and Affinity Purification System

The NativePure™ Lentiviral Expression and Affinity Purification System provides an efficient method for lentiviral delivery of your gene of interest into cells, and the purification and analysis of native protein complexes. The NativePure™ Lentiviral Gateway® Vectors allow N- or C- terminal fusion of your recombinant protein of interest to the capTEV± affinity purification tag.

The Lentiviral Expression and Affinity Purification System provides:
• Gateway® vectors designed to express your protein of interest in multiple expression systems
• Lentiviral vectors with the capTEV™ Tag that allow in vivo biotinylation of your protein of interest
• Streamlined purification protocols with Streptavidin Agarose

Contents:
The NativePure™ Lentiviral Expression and Affinity Purification System includes the NativePure™ Lentiviral Gateway® Vector Kit, the NativePure™ Lentiviral Expression Kit, and the NativePure™ Lentiviral Affinity Purification Kit. Each kit is also available separately.

The NativePure™ Lentiviral Gateway® Vector Kit consists of the pLenti6/capTEV™-NT-DEST1 (N-terminal vector), pLenti6/capTEV™-CT-DEST (C-terminal vector), and pLenti6/capTEV™-CT-GW/ARP2 (C-terminal) control vector.

The NativePure™ Lentiviral Expression Kit contains the NativePure™ Lentiviral Gateway® Vector Kit, the ViraPower™ Bsd Lentiviral Support Kit, 293FT Cell Line, One Shot® Stbl3™ Chemically Competent E. coli, and Gateway® LR Clonase® Enzyme Mix.
The NativePure™ Lentiviral Affinity Purification Kit contains the NativePure™ Lentiviral Gateway® Vector Kit, Streptavidin Agarose, NativePure™ 5X Lysis/Binding Buffer, NativePure™ 10X TEV Cleavage Buffer, 100 mM DTT (Dithiothreitol), 10% NP40, AcTEV™ Protease, NativePure™ Columns, and NativePure™ Concentrators.

Store the 293FT Cell Line in liquid nitrogen and the One Shot® Stbl3™ Chemically Competent E. coli at -80°C.

Store Streptavidin Agarose, 10% NP-40, NativePure± 5X Lysis/Binding Buffer, NativePure± 10X TEV Cleavage Buffer, Lipofectamine™ 2000, NativePure± Columns, and NativePure± Concentrators at +4°C.

Store the NativePure™ Lentiviral Gateway® Vector Kit, AcTEV± Protease, 100 mM DTT, Gateway® LR Clonase® Enzyme Mix, ViraPower™ packaging Mix, and Blasticidin at -20°C. All components are guaranteed stable for 6 months when properly stored.

pBAD/TOPO™ ThioFusion™ Expression Kit (Invitrogen™)

The pBAD/TOPO® ThioFusion™ Expression Kit is designed for one-step cloning and regulated expression of thioredoxin fusion proteins in E. coli(Figure 1). The pBAD/Thio-TOPO® vector encodes His-Patch thioredoxin as an N-terminal fusion partner. The thioredoxin fusion can significantly increase the solubility of many difficult-to-express proteins and improve the yield of protein production. The vector includes the following additional features (Figure 2):

• The araBAD promoter for tightly regulated expression in E. coli I-activated vector for 5-minute TOPO® Cloning of Taq-amplified PCR products
• C-terminal polyhistidine (6xHis) tag for purification with nickel-chelating resin and detection with an Anti-His(C-term) Antibody
• C-terminal V5 epitope for detection with an Anti-V5 Antibody
• Enterokinase cleavage site for efficient cleavage of N-terminal fusion tag with EKMax™ Enterokinase

Champion™ pET161 Directional TOPO™ Expression Kit with Lumio™ Technology (Invitrogen™)

The Champion™ pET Expression System yields the highest-level protein production in E. coli. During expression, your protein of interest can reach levels greater than 50 percent of total cellular protein. Based on T7 expression vectors originally developed by Studier and colleagues (1-3), high-level expression is achieved because the T7 RNA polymerase is more processive than native E. coliRNA polymerase and is dedicated to the transcription of your gene of interest. Protein production is further enhanced in the system by the expression strain BL21 Star™ E. coli, which significantly improves the stability of mRNA transcripts and increases protein expression up to ten-fold.

Simplified, efficient Directional TOPO® cloning allows you to quickly enter a Champion™ pET Expression vector. These kits feature linearized, topoisomerase I-activated Champion™ pET expression vectors for five-minute directional cloning. Directional TOPO® Cloning technology facilitates gene expression because:

• A proofreading enzyme is used for PCR, resulting in fewer errors in cloned genes
• Greater than 90% of the clones are in the correct orientation for gene expression, reducing the time spent on colony screening

Seven Champion™ pET Directional TOPO® Expression Vectors are available (Figure 1 and Table 1): Each vector carries a T7lac promoter for high-level expression. Flexible options for simplifying protein detection, cleaving purification tags, selecting plasmid carrying clones, and/or improving protein yields are available.
With the Champion™ pET Directional TOPO® vectors you can expect highest-level protein production. Figure 2 shows expression of the lacZ gene in Champion™ pET Directional TOPO® vectors. Figure 3 demonstrates efficient cleavage using TEV protease of the N-terminal tag of a β-galactosidase fusion protein expressed from pET151/D-TOPO®.

Bac-to-Bac™ C-His TOPO™ Expression System (Gibco™)

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 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® C-His TOPO® cloning or expression kit, you can produce a C-terminal His-fusion protein with a TEV cleavage site to purify with nickel-chelating resins (Invitrogen’s ProBond™ Purification System) and generate a native protein with the aid of Invitrogen’s popular AcTEV™ Protease. This is the Invitrogen’s only vector that contains a C-terminal TEV cleavage site!

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.

ExpiCHO™ Expression System Kit (Gibco™)

The ExpiCHO™ Expression System Kit combines the power of rapid, ultra–high-yield transient protein production in suspension culture with the benefit of expression in Chinese hamster ovary (CHO) cells. It is based on high-density culture of ExpiCHO-S™ cells in ExpiCHO™ Expression Medium. Transient expression is powered by ExpiFectamine™ CHO transfection reagent, designed specifically for high-density CHO suspension culture, in combination with a specialized transfection enhancer and culture feed. All components work together synergistically to generate 2- to 10-fold higher protein yields than the HEK293-based Expi293™ Expression System Kit and 20- to 150-fold higher yields than the FreeStyle™ MAX CHO Expression System. Expression yields of 1 to 3 g per L of transfected culture have been demonstrated for some antibody and non-antibody proteins.

The ExpiCHO Expression System offers:

• Milligram to multigram yields of recombinant protein per liter of culture in 5 to 14 days post-transfection, which are frequently higher than transient 293-based systems
• Cost-effective transient expression in CHO cells, allowing scientists developing protein biotherapeutics to work with CHO-expressed proteins from start to finish in the drug development process
• A robust alternative mammalian transient expression host, allowing for production of proteins that are difficult to express in HEK293 cells
• Flexible, reliable culture and transfection protocols with multiple options that readily fit into current transient expression workflows
• Scalable expression for cultures from <1 mL to >10 L, thus generating the amount of protein needed for your application

The components included in the ExpiCHO Expression System Kit are: 2 vials of frozen ExpiCHO-S cells, 1 L of ExpiCHO Expression Medium, 1 ExpiFectamine CHO Transfection Kit sufficient to transfect 1 L of culture, OptiPRO™ SFM, and an antibody-expressing positive control vector. All components of the ExpiCHO Expression System Kit are also available for purchase separately. All components of the system are animal origin–free.

Jump-In™ CHO-K1 Kit

The Jump-In™ CHO-K1 Kit allows the targeted integration of genetic material into a specific pre-engineered R4 site in the Jump-In™ CHO-K1 cell line, to create an isogenic stable cell line with less effort and in less time than traditional cell engineering methods.

The high retargeting efficiency, made possible by the R4 sites in the CHO-K1 cell line, allows the use of the isogenic pool for additional experiments without the need for clonal selection. Alternatively, the high retargeting efficiency allows for the easy selection of a positive stable clone expressing your gene of interest.

The Jump-In™ CHO-K1 Kit Lets You:

• Quickly and efficiently develop stably engineered isogenic cell pools in about half the time compared to traditional cell engineering methods
• Utilize isogenic expression from a defined genomic locus as the ideal solution for comparative analysis of gene families, isoforms, or orthologs
• Generate multiple cell lines in parallel using the simplified work flow
• Easily access the technology without complicated licenses or restrictions to interpret

Save Time with Rapid and Efficient Generation of Engineered Cell Lines
With the Jump-In™ CHO-K1 Kit you can generate functional cell pools in as little as 2 weeks without laborious clone isolation and analysis, and the streamlined protocol makes it easier to generate several cell lines at the same time. Even generation of clonal cell lines can be done with reduced time and effort due to the high percentage of positive clones. In addition, the Jump-In™ technology gives you the freedom to generate an unlimited number of cell lines without restrictive licensing requirements.

Expand Your Experimental Capabilities
The Jump-In™ CHO-K1 Kit is the ideal solution for cells and assays where transient engineering technologies are problematic, as well as for difficult to The kit also provides a convenient way to create target panels of gene families, isoforms, or orthologs. Genes coding for large proteins or multi-unit proteins are not a problem since the Gateway® destination vectors accept large inserts.

The kit includes:
• pJTI™ R4 Dest CMV pA vector (100 µg)
• pJTI™ R4 Int vector (100 µg)
• Jump-In™ CHO-K1 Cells (2 vials @ 1 ml each)

For Research Use Only. Not intended for any animal or human therapeutic or diagnostic use.

1-Step CHO High-Yield IVT Kit (Thermo Scientific™)

The Thermo Scientific™ 1-Step CHO High-Yield In Vitro Translation (IVT) Kit enables the expression of functional proteins with up to 750 times greater yield per mL of reaction than rabbit reticulocyte lysate-based mammalian IVT systems that only yield up to 10 μg protein/mL reaction.

• High expression—up to 750 µg/mL of expressed protein
• Reproducible results—low variability between experiments
• Fast—express high levels of protein with 6 hours to overnight incubation
• Easier storage—allows you to store lyophilized CHO cell lysate at -20°C
• Functional—obtain functionally active proteins, including those containing disulfide bonds
• Easy-to-use—three reaction components together carry out transcription and translation in one step
• Great for labeling—amenable to incorporation of labeled and unnatural amino acids
• Vector compatibility—uses existing pT7CFE1 vectors for protein expression

The 1-Step CHO High-Yield IVT System uses extracts containing the robust translation machinery of the pharmaceutical industry standard Chinese Hamster Ovary (CHO) cell line to generate functional full-length proteins. In this system, protein expression is performed in a proprietary dialysis device that allows a continuous supply of nucleotides, amino acids, and energy-generating substrates into the reaction while removing inhibitors of proteins synthesis. This continuous-exchange cell-free (CECF) system enables protein expression in an overnight incubation of up to 750 µg/mL. The CHO IVT kit includes all of the components required for transcription and translation of a recombinant gene, including an optimized expression vector.

Includes:
Lyophilized cell lysate, reaction mix, accessory proteins, 4X dialysis buffer, control DNA, cloning vector, dialysis devices, and centrifuge tubes.

Requires:
30°C incubator or water bath (shaker incubator increases expression by 30 to 50%)

Applications:
• Production of large amounts of recombinant protein for structural analysis
• Generation of active proteins for downstream applications such as protein interaction studies
• Rapid large scale expression and characterization of mutant proteins
• Production of viral particles
• Incorporation of unnatural amino acids
• Production of cytotoxic proteins

MembraneMax™ Protein Expression Kit (Invitrogen™)

MembraneMax™ Protein Expression Kits delivers high yields of soluble (dispersed) membrane proteins using the MembraneMax™ reagent, which is an planar phospholipid membrane bilayer surrounded by a scaffold protein (also called a nanolipoprotein particle or NLP). Combining this technology with a scalable cell-free expression E. coli extract delivers microgram to milligram quantities of your membrane protein specifically for high throughput expression screening, expression of membrane proteins that would otherwise be toxic in cell-based systems and functional or activity assays. With high uniformity and consistent structure, MembraneMax reagent minimizes formation of membrane protein aggregates or clumping often caused by inconsistent size, shape and structure typical observed with microsomes or micelles. The MembraneMax™ Protein Expression kits contain all the necessary reagents for successful expression of your membrane protein.

Key Advantages of the MembraneMax™ Protein Expression Kits:
• Optimized cell-free expression delivers high yield of membrane protein from nanogram to milligram quantities
• MembraneMax™ reagent enables a monodispersed population of soluble membrane protein-NLP complexes
• Expression format amenable to high-throughput protein synthesis for screening and expression of toxic proteins
• Production of a homogeneous population of your membrane protein
• Convenient kit format includes the necessary reagents for protein expression—simply add your gene of interest to get started

1-Step Human High-Yield Mini IVT Kit (Thermo Scientific™)

The Thermo Scientific 1-Step Human High-Yield In Vitro Translation (IVT) Kits enable the expression of functional proteins with 10 to 100 times greater yield per mL than other mammalian IVT systems.

The 1-Step High-Yield IVT System uses modified HeLa cell extracts to take advantage of the robust human translation machinery and generate functional full-length proteins. In this system, protein expression is performed in a proprietary dialysis device that allows a continuous supply of nucleotides, amino acids and energy generating substrates into the reaction while removing inhibitors of proteins synthesis. This continuous-exchange cell-free (CECF) system enables protein expression in an overnight incubation of up to 750 µg/mL. The complete mini-scale kit includes all the components required for transcription and translation of a recombinant gene, including an optimized expression vector.

Features of the 1-Step Human High-Yield Mini IVT Kit:

High expression—up to 750 µg/mL of expressed protein
Reproducibility—low variability between experiments
Fast—express high levels of protein with 6 hours to overnight incubation
Functional—obtain functionally active proteins, including those containing disulfide bonds

1-Step Protein Expression:
Easy-to-use—transcription and translation is performed in one reaction step
Adapt for labeling—amenable to incorporation of heavy or unnatural amino acids

Includes:
• Complete kits contain cell lysate, reaction mix, accessory proteins, 5X dialysis buffer, control DNA, cloning vector, dialysis devices and centrifuge tubes

Requires:
• 30°C incubator or water bath (shaker incubator increases expression by 30 to 50%)

Applications:
• Production of recombinant proteins for structural analysis
• Generate active proteins for protein interaction studies
• Perform rapid expression and characterization of mutant proteins
• Small-scale production of viral particles
• Incorporation of unnatural amino acids or toxic protein production

The 1-Step Human High-Yield IVT Kits are cell-free protein expression systems that provide all of the essential components required for transcription and translation. The kits are optimized with Accessory Proteins and Reaction Mixes that support protein synthesis using a DNA template. The advantages of using the 1-Step Human High-Yield IVT Kits over traditional in vivo expression systems include the ability to express toxic or insoluble proteins, easily perform protein labeling with modified amino-acids and reduce the time and cost of expressing human proteins in tissue culture cells. 1-Step Human IVT Kits are optimized for use with the pT7CFE1 expression vector that utilizes the T7 viral promoter and an EMCV Internal Ribosome Entry Site (IRES) to facilitate high levels of in vitro protein expression. Using a pT7CFE vector is critical for obtaining high expression levels. For added convenience, at family of pT7CFE vectors are available with tandem affinity tags for facilitate protein detection and purification. (For smaller scale and screening options, consider the 1-Step Human Coupled IVT Kit.)

More Product Data
Choosing a vector and purification method for in vitro protein expression
System for high-throughput, high-yield in vitro translation
Expression of highly active proteins using a cell-free human IVT system

Multi-Copy Pichia Expression Kit (Invitrogen™)

The Multi-Copy Pichia Expression Kit contains the pPIC3.5K, pPIC9K, and pAO815 vectors for production and selection of Pichia strains that contain more than one copy of the gene of interest. In many cases, increased copy number has led to increased expression levels.

pPIC9K and pPIC3.5K
The pPIC9K and pPIC3.5K vectors carry the kanamycin resistance gene that confers resistance to Geneticin® Reagent in Pichia. Spontaneous generation of multiple insertion events can be identified by resistance to increased levels of Geneticin® Reagent. Pichia transformants are selected on histidine-deficient medium and screened for their level of resistance to Geneticin® Reagent. The ability to grow in high concentrations of Geneticin® indicates that multiple copies of the kanamycin resistance gene and the gene of interest are integrated into the genome (1). The pPIC9K vector directs secretion of expressed proteins while proteins expressed from pPIC3.5K remain intracellular.

pAO815
pAO815 is a Pichia expression vector designed to clone multiple copies of a gene into a single vector. The vector contains a Bgl II site upstream of the 5´ AOX1 gene and a unique BamH I site downstream of the 3´ AOX1 transcription termination (TT) signal. Four steps are required to generate multiple copies of the gene of interest:
1. The gene is cloned into the unique EcoR I site in the vector.
2. The construct is digested with BamH I and Bgl II to release the "expression cassette" containing the AOX1 promoter, gene of interest, and 3´ AOX1 TT.
3. Concatemers of the expression cassette are generated by ligation in vitro.
4. The multiple copies are inserted back into the pAO815 vector and transformed into Pichia.

These vectors are also available separately.

Bac-to-Bac™ N-His TOPO™ Expression System (Gibco™)

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.