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Gateway™ pDEST™24 Vector (Invitrogen™)

To fit all of your expression needs, Invitrogen offers state-of-the-art Gateway® destination vectors for expression in E. coli, insect, yeast, or mammalian cells, as well as for production of native protein or N- or C-terminal fusion proteins. All Gateway® destination vectors have attR sites for recombination with any attL-flanked fragment, regardless of whether it is an entry clone or an Ultimate™ RF Clone. The following table lists the wide range of destination vectors available.

Additional materials required, available separately: Gateway® entry clone, Gateway® LR Clonase® enzyme mix, and reaction buffer.

pZeoSV2(+) Vector Kit (Invitrogen™)

pZeoSV2(+) and (-) are 3.5-kb vectors designed for high-level, constitutive expression in mammalian cell lines that express the SV40 large T antigen. The Zeocin™ resistance gene provides fast and efficient selection in E. coliand mammalian cells that do not express the SV40 large T antigen. The vectors have the following features which make them efficient, easy-to-use tools:

• SV40 enhancer-promoter and origin for high-level, constitutive expression and replication in mammalian cells
• BGH polyA signal for efficient processing of mRNA transcripts
• Multiple cloning site in the forward (+) and reverse (-) orientations for easier cloning
• Small size for efficient cloning
• f1 origin for the rescue of single-strand DNA (sense strand)
• pUC origin of replication for growth in E. coli

Expression of the Zeocin™ resistance gene is driven by the CMV promoter in mammalian cell lines and by the synthetic EM-7 promoter in E. coli. pZeoSV2 can be used in transient expression assays as well as to create stable cell lines.

Champion™ pET302/NT-His and pET303/CT-His Vector Kit (Invitrogen™)

The Champion™ pET302/NT-His and pET303/CT-His Vector Kit is designed for cloning a gene of interest via restriction enzyme(s) and ligase (REaL) and subsequent high-level expression from the strong bacteriophage T7 promoter. In addition to the T7 promoter, each vector contains only the necessary functional elements and an N- or C-terminal 6xHis tag (pET302/NT-His and pET303/CT-His, respectively) for convenient purification and detection (Figure 1). Expression levels obtained from these vectors may be higher than those obtained from another suppliers pET vector (Figure 2). To maximize expression, use with MagicMedia™ E. coli Expression Medium.


Contents and Storage:

The Champion™ pET302/NT-His and pET303/CT-His Vector Kit includes 6 µg each of pET302/NT-His and pET303/CT-His and 10 µg of a control vector. Store at -20“C. All components are guaranteed stable for 6 months when properly stored.

pLenti7.3/V5-DEST™ Gateway™ Vector Kit (Invitrogen™)

The pLenti7.3⁄V5-DEST™ Gateway® Vector Kit contains the Gateway®-adapted ViraPower™ HiPerform™ lentiviral expression vector, pLenti7.3⁄V5-DEST™ for easy recombination-based cloning and high-level expression of a target gene in dividing and non-dividing mammalian cells. The pLenti7.3⁄V5-DEST™ vector is equipped with two key genetic elements, making it a HiPerform™ vector: the Woodchuck Posttranscriptional Regulatory Element (WPRE) and the central Polypurine Tract (cPPT) sequence from the HIV-1 integrase gene to produce at least 4-fold increase in protein expression compared to vectors lacking these elements. In addition, the vector allows for an accurate determination of titer of functional lentivirus in just two days using Emerald Green Fluorescent Protein (EmGFP).

Advantages
• Transient expression
• Short-term experiments
• High-throughput screening
• 2 day titer of functional virus using EmGFP
• Flexible and versatile Gateway® recombination cloning technology

Key Features
• WPRE (Woodchuck Posttranscriptional Regulatory Element) from the woodchuck hepatitis virus, allows increased transgene expression and cPPT (Polypurine Tract) from the HIV-1 integrase gene, increases the copy number of lentivirus integrating into the host genome
• Human cytomegalovirus (CMV) immediate early promoter to control high-level expression of the gene of interest
• SV40 promoter driving expression of EmGFP
• Emerald Green Fluorescent Protein (EmGFP) allows easy determination of Lentiviral titers by flow cytometry and monitor transduced cells.

Kit Includes
• pLenti7.3⁄V5-DEST™ Gateway® Vector box
• One Shot® Stbl3™ chemically competent E. coli (C7373-03)

Related SKUs
• 293FT Cell Line (R70007; R70007)
• ViraPower™ Lentiviral Support Kit (K4970-00)
• ViraPower™ Lentiviral Packaging Mix (K4975-00)
• Lipofectamine® 2000 (11668-019; 11668-027)

For research use only. Not intended for any therapeutic or diagnostic use.

pcDNA™3.1/nV5-DEST Mammalian Expression Vector (Invitrogen™)

To fit all of your expression needs, Invitrogen offers state-of-the-art Gateway® destination vectors for expression in E. coli, insect, yeast, or mammalian cells, as well as for production of native protein or N- or C-terminal fusion proteins. All Gateway® destination vectors have attR sites for recombination with any attL-flanked fragment, regardless of whether it is an entry clone or an Ultimate™ RF Clone. The following table lists the wide range of destination vectors available.

Additional materials required, available separately: Gateway® entry clone, Gateway® LR Clonase® enzyme mix, and reaction buffer.

pT7CFE1-NFtag Vector for Mammalian Cell-Free Protein Expression (Thermo Scientific™)

Thermo Scientific pT7CFE1-NFtag is a cloning plasmid optimized to use with the Thermo Scientific 1-Step Human In Vitro Protein Expression System for in vitro translation (IVT) of tagged fusion proteins. pT7CFE1-NFtag Vector is available with single affinity Ftag tag at the N-terminus to facilitate protein purification and detection.

Features of pT7CFE1:
• EMCV IRES at the 5' UTR promotes high-level translation of mRNAs
• MCS accommodates gene insertion via ten different restriction sites: Msc1, Nde1, BamH1, EcoR1, EcoRV, Pac1, Pst1, Sac1, Sal1, Not1 and Xho1
• Poly A sequence in the 3' region promotes mRNA stabilization and protection from nucleases
• T7 terminator ensures synthesis of accurate size mRNA transcripts
• Plasmid linearization may be accomplished with restriction sites between Poly A sequence and the T7 terminator region

pT7CFE1 Expression Vectors contain the Encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) element that is critical for high levels of cap-independent protein expression in the Human In Vitro Translation System. Each vector features a highly-compatible multiple cloning site (MCS) to facilitate easy insertion of protein coding sequences into and between vectors. The pT7CFE1 Vector is available with single or tandem affinity tags at the N- or C- terminus to facilitate protein purification and detection. The pT7CFE Vectors are suitable for insertion of cloned genes, cDNAs, ORFs or PCR products for in vitro transcription and translation. Custom cloning services are also available.

More Product Data
Choosing a vector and purification method for in vitro protein expression

Related Products
pT7CFE1-Cftag Vector for Mammalian Cell-Free Protein Expression
pT7CFE1-CGST-HA-His Vector for Mammalian Cell-Free Protein Expression
pT7CFE1-NHis-GST Vector for Mammalian Cell-Free Protein Expression
pT7CFE1-NHis-GST-CHA Vector for Mammalian Cell-Free Protein Expression

pLenti6/V5-DEST™ Gateway™ Vector (Invitrogen™)

The pLenti6⁄V5-DEST® Gateway® Vector is a Gateway®-adapted ViraPower™ lentiviral expression vector for lentiviral-based expression of a target gene in dividing and non-dividing mammalian cells. The vector has the CMV promoter for driving constitutive expression of the target gene and the blasticidin selection marker for stable selection in mammalian cells.

Advantages
• Lentivirus based expression of a target gene in dividing and non-dividing mammalian cells

Key Features
• Flexible and versatile Gateway® recombination cloning technology
• Constitutive high expression with CMV promoter
• Blasticidin selection marker for stable selection
• C terminal V5 tag for quick detection

Kit includes
• pLenti6⁄V5-DEST™ Gateway® Vector
• One Shot® Stbl3™ Chemically Competent E. coli (C7373-03)

Related SKUs
• pLenti6⁄UbC⁄V5-DEST™ Gateway® Vector (V49910)
• pLenti4⁄V5-DEST™ Gateway® Vector (V49810)
• pLenti6⁄V5 Directional TOPO® Cloning Kit (K4955-10)
• ViraPower™ Lentiviral Directional TOPO® Expression Kit (K495000)
• ViraPower™ Lentiviral Gateway® Expression Kit (K4960-00)
• ViraPower™ HiPerform™ Lentiviral Gateway® Expression Kit (K5330-00)

For research use only. Not intended for any therapeutic or diagnostic use.

pcDNA™3.1 (-) Mammalian Expression Vector (Invitrogen™)

This pcDNA™3.1(-) vector is designed for high-level, constitutive expression in a variety of mammalian cell lines. It contains a Geneticin® selectable marker and a reverse-orientation multiple cloning site.

The pcDNA™3.1 Expression Vector Family
Three untagged versions of pcDNA™3.1 (available separately), each with a different selectable marker (Geneticin®, Zeocin™, or Hygromycin), are for use alone or in co-transfections. All three vectors offer the following features:
• Cytomegalovirus (CMV) enhancer-promoter for high-level expression
• Large multiple cloning site in either forward (+) or reverse (-) orientations
• Bovine Growth Hormone (BGH) polyadenylation signal and transcription termination sequence for enhanced mRNA stability
• SV40 origin for episomal replication and simple vector rescue in cell lines expressing the large T antigen (i.e., COS-1 and COS-7)
• Ampicillin resistance gene and pUC origin for selection and maintenance in E. coli

pVAX1 Vector (Invitrogen™)

pVAX1 is specifically designed for use in the development of DNA vaccines. pVAX1 has the following features:

• Eukaryotic DNA sequences limited to those required for expression in order to minimize the possibility of chromosomal integration
• Kanamycin resistance gene for selection in E. colibecause ampicillin has been reported to cause an allergic response in some individuals
• Expression levels of recombinant proteins from pVAX1 comparable to those achieved with its parent vector, pcDNA™3.1

The small size of pVAX1 and the variety of unique cloning sites simplify subcloning of even very large DNA fragments.

MultiSite Gateway™ Pro Plus, for flexible cloning of up to four DNA fragments into a Gateway™ destination vector (Invitrogen™)

MultiSite Gateway® Pro Technology enables you to efficiently and conveniently assemble multiple DNA fragments in the desired order and orientation into a Gateway® Expression vector. Using specifically designed att sites for recombinational cloning, you can clone two, three, or four DNA fragments into any Gateway® Destination vector containing attR1 and attR2 sites. The resulting expression clone is ready for downstream expression and analysis applications. The new MultiSite Gateway® Pro Technology allows you to:


• Clone multiple DNA fragments into one vector without using restriction enzymes or ligases
• Take full advantage of the wide selection of Gateway® Destination vectors (available from Invitrogen) or create your own
• Replace multiple plasmid transfections with a single vector carrying all DNA elements of interest


MultiSite Gateway® Pro Kits are available for cloning two, three, or four DNA fragments and for flexible and customized configuration of DNA fragments into a Gateway® Destination vector. All of the kits include internal positive control entry clones for troubleshooting potential entry clone issues.

BLOCK-iT™ Lentiviral RNAi Zeo Gateway™ Vector Kit (Invitrogen™)

The BLOCK-iT™ Lentiviral RNAi Zeo Gateway® Vector Kit contains the pLenti4/BLOCK-iT™-DEST expression vector which enables lentiviral delivery and genomic integration of DNA coding for shRNA. Once expressed, the shRNA is processed by cellular machinery and initiates target-specific RNAi. The pLenti4/BLOCK-iT™-DEST vector offers:

Gateway® Technology for efficient recombination of the RNAi cassette from the BLOCK-iT™ inducible pENTR™/H1/TO vector

• All required components for efficient lentiviral packaging, delivery, and integration of the shRNA
• Zeocin™ selection marker for fast selection of clonal cell lines containing the RNAi cassette

pcDNA™5/FRT/TO Vector Kit (Invitrogen™)

pcDNA™FRT⁄TO vector is a 5.1 kb inducible expression vector designed for use with the Flp-In™ T-REx™ System. The control plasmid, pcDNA™5⁄FRT⁄TO⁄CAT, is also included for use as a positive control for transfection and expression in your Flp-In™ T-REx™ host cell line of choice.

The pcDNA™5⁄FRT⁄TO vector contains the following elements:

• A hybrid human cytomegalovirus (CMV)⁄TetO2 promoter for high-level, tetracycline-regulated expression of the gene of interest in a wide range of mammalian cells
• Multiple cloning site with 10 unique restriction sites to facilitate cloning the gene of interest
• FLP Recombination Target (FRT) site for Flp recombinase-mediated integration of the vector into the Flp-In™ T-REx™ host cell line
• Hygromycin resistance gene for selection of stable cell lines

pShooter™ Mammalian Expression Vector (pCMV/myc/cyto) (Invitrogen™)

The pShooter™ vectors are designed to localize recombinant proteins to the nucleus, mitochondria, endoplasmic reticulum, or cytoplasm. In addition to the localization signal, each pShooter™ vector contains:

• Strong mammalian promoter (CMV or EF-1É) for constitutive expression of your protein of interest
• C-terminal c-myc epitope for detection with an Anti-myc Antibody
• Bovine Growth Hormone (BGH) polyadenylation site for mRNA stability
• f1 origin for single-stranded rescue of sense DNA
• Neomycin resistance gene for selection of mammalian cells with Geneticin¤ selection agent
• Ampicillin resistance and pUC origin for selection and maintenance in E. coli

Each vector is provided with a positive control plasmid. The positive control expresses GFP and targets it to the same subcellular location as the pShooter™ vector.

PichiaPink™ Secreted Vector Kit (Invitrogen™)

The PichiaPink™ Secreted Vector Kit contains the pPINKα-HC vector for use with the PichiaPink™ Yeast Expression System. The pPINKα-HC vector is built around the ADE2 gene for complementing the complementing the ade2 deletion in the PichiaPink™ strains. The pPINKα-HC vector has the α-mating factor secretion signal sequence so that all proteins are secreted. You can order the PichiaPink™ Secreted Vector Kit to refill your PichiaPink™ Yeast Expression System Kit.

The PichiaPink™ Secreted Vector Kit comes with:
•     pPINKα-HC vector (high-copy number) (20 µg)
•     5’α-factor sequencing primer (20 µg)
•     3’CYC1 sequencing primer (20 µg)
•     One Shot® TOP10™ Electrocomp™ E. coli (Cat. No. C404052)

Why Choose the PichiaPink™ Yeast Expression System?
The PichiaPink™ Yeast Expression System is based on the yeast Pichia pastoris. Advantages of Pichia pastoris include rapid growth, well-defined genetic background, simple media formulation, and easy handling. For over 30 years, Pichia pastoris has been used by labs around the world for producing hundreds of different proteins from many species including human (Ref 1, 2). The PichiaPink™ Yeast Expression System allows convenient and cost-effective protein production from small to large scales.

For information on obtaining a commercial-use license for the PichiaPink™Yeast Expression System, please inquire at outlicensing@lifetech.com.

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

Related Links
Download the pPINKα-HC plasmid map (PDF).
Learn more about the PichiaPink™ Yeast Expression System.
Learn more about our other protein expression systems.

References

1. Cereghino JL, Cregg JM. Heterologous protein expression in the methylotrophic yeast Pichia pastoris. FEMS Microbiol Rev. 2000 Jan;24(1):45-66. [PubMed]
2. Cereghino GP, Cereghino JL, Ilgen C, Cregg JM. Production of recombinant proteins in fermenter cultures of the yeast Pichia pastoris. Curr Opin Biotechnol. 2002 Aug;13(4):329-32. [PubMed]

pT7CFE1-CHis Vector for Mammalian Cell-Free Protein Expression (Thermo Scientific™)

Thermo Scientific pT7CFE1-CHis is a cloning plasmid optimized to use with the Thermo Scientific 1-Step Human In Vitro Protein Expression System for in vitro translation (IVT) of tagged fusion proteins. pT7CFE1-CHis Vector is available with single affinity 6xHis tag at the C-terminus to facilitate protein purification and detection.

Features of pT7CFE1:
• EMCV IRES at the 5' UTR promotes high-level translation of mRNAs
• MCS accommodates gene insertion via ten different restriction sites: Msc1, Nde1, BamH1, EcoR1, EcoRV, Pac1, Pst1, Sac1, Sal1, Not1 and Xho1
• Poly A sequence in the 3' region promotes mRNA stabilization and protection from nucleases
• T7 terminator ensures synthesis of accurate size mRNA transcripts
• Plasmid linearization may be accomplished with restriction sites between Poly A sequence and the T7 terminator region

pT7CFE1 Expression Vectors contain the Encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) element that is critical for high levels of cap-independent protein expression in the Human In Vitro Translation System. Each vector features a highly-compatible multiple cloning site (MCS) to facilitate easy insertion of protein coding sequences into and between vectors. The pT7CFE1 Vector is available with single or tandem affinity tags at the N- or C- terminus to facilitate protein purification and detection. The pT7CFE Vectors are suitable for insertion of cloned genes, cDNAs, ORFs or PCR products for in vitro transcription and translation. Custom cloning services are also available.

More Product Data
Choosing a vector and purification method for in vitro protein expression

Related Products
pT7CFE1-NHis Vector for Mammalian Cell-Free Protein Expression
pT7CFE1-CGST-HA-His Vector for Mammalian Cell-Free Protein Expression
pT7CFE1-NHis-GST Vector for Mammalian Cell-Free Protein Expression
pT7CFE1-NHis-GST-CHA Vector for Mammalian Cell-Free Protein Expression

pJTI™ R4 CMV-TO MCS pA Vector

The pJTI™ R4 CMV-TO MCS pA vector is designed for the expression of your gene of interest under the control of the tet-inducible CMV promoter after restriction enzyme cloning and retargeting into the genomic R4 site of a Jump-In™ parental cell line.

This vector can be used for inducible or constitutive expression of your gene of interest, depending on which Jump-In™ parental cell line you use. When this vector is retargeted into a Jump-In™ T-REx™ parental cell line, gene expression is controlled by the tet-operon and can be induced by adding doxycycline to the growth media. When used with a Jump-In™ parental cell line such as the Jump-In™ GripTite™ HEK293 cell line, constitutive expression is achieved after retargeting. The R4 sites in the Jump-In™ parental cell lines result in a high retargeting efficiency, requiring less effort and less time than traditional cell engineering methods. Retargeting of Jump-In™ parental cell lines results in creation of an isogenic pool that is sufficient for cell-based experiments without the need for clonal selection. Alternatively, the high retargeting efficiency allows for easy selection of a positive stable clone for expressing your gene of interest.

TA Cloning™ Kit, Dual Promoter, with pCR™II Vector and One Shot™ INVαF' Chemically Competent E. coli (Invitrogen™)

The TA Cloning® Kit Dual Promoter with pCR™II vector provides a quick, one-step cloning strategy for directly inserting a Taq-amplified PCR product into a plasmid vector. The T7 and Sp6 promoters of the pCR™II vector allow in vitro transcription of the insert to produce sense or anti-sense products. The TA Cloning Kit Dual Promoter uses the pCR™II cloning vector and ExpressLink™ T4 DNA Ligase to generate a ligation product in a fifteen-minute, room-temperature ligation step. Reactions typically yield >80% recombinants containing inserts.

Features of the TA Cloning® Kit Dual Promoter:
Fast & convenient—15-minute, room-temperature ligation
Efficient—blue/white screening and >80% clones with correct insert
Flexible—choice of kanamycin or ampicillin resistance for flexible antibiotic selection
Hassle-free—eliminates any enzymatic modifications of the PCR product
Streamlined—does not require the use of PCR primers that contain restriction sites

The pCR™II vector provides:
• 3'-T overhangs for direct ligation of Taq-amplified PCR products
• T7 and Sp6 promoters for in vitro RNA transcription and sequencing
• Versatile polylinker with flanking EcoR I sites for easy excision of inserts
• M13 forward and reverse primer sites for sequencing

How TA Cloning® Works
Taq polymerase has a non-template-dependent activity that adds a single deoxyadenosine (A) to the 3' ends of PCR products. The linearized vector supplied in this kit has single 3' deoxythymidine (T) residues. This allows PCR inserts to ligate efficiently with the vector.

Kit Configurations
The TA Cloning® Kit is offered in a variety of configurations: with One Shot® INVF' Chemically Competent E. coli (K2050-01 and K2050-40), One Shot® TOP10F' Chemically Competent E. coli (K2060-01 and K2060-40), and without competent cells (K2750-20 and K2750-40) in 20- and 40-reaction kit sizes.

TOPO™ TA Cloning™ Kit, Dual Promoter, with pCR™II-TOPO™ Vector and One Shot™ TOP10 Electrocomp™ E. coli (Invitrogen™)

TOPO® TA Cloning® Kits are designed for cloning PCR products directly from a PCR reaction in just 5 minutes (1). They use a pCR™-TOPO® Vector with covalently bound topoisomerase I for fast cloning and recombinants. pCR™-TOPO® Vectors include:

• 3´-T overhangs for direct ligation of Taq-amplified PCR products
• Choice of T7 (pCR™2.1-TOPO®) or T7 and SP6 (pCR™II-TOPO®) promoters for in vitro RNA transcription and sequencing. Each vector also contains M13 forward and reverse primer sites for sequencing.
EcoR I sites flanking the PCR product insertion site for easy excision of inserts
• Kanamycin and ampicillin resistance genes for your choice of selection in E. coli
• Easy blue/white colony screening for selection of recombinants

TOPO® TA Cloning® Kits are also available in combo kits combined with a PureLink™ Quick Plasmid Miniprep Kit (50 preps) for fast plasmid purification of your TOPO®-cloned inserts for downstream analysis.

TA Cloning™ Kit, with pCR™2.1 Vector and One Shot™ TOP10F' Chemically Competent E. coli (Invitrogen™)

The TA Cloning® Kit with pCR™2.1 vector provides a quick, one-step cloning strategy for directly inserting a Taq-amplified PCR product into a plasmid vector. The TA Cloning® Kit uses the pCR™2.1 cloning vector and ExpressLink™ T4 DNA Ligase to generate a ligation product in a fifteen-minute, room-temperature ligation step. Reactions typically yield >80% recombinants containing inserts.

Features of the TA Cloning® Kit with pCR™2.1 vector:
Fast & convenient—15-minute, room-temperature ligation
Efficient—blue/white screening and >80% clones with correct insert
Flexible—choice of kanamycin or ampicillin resistance for flexible antibiotic selection
Hassle-free—eliminates any enzymatic modifications of the PCR product
Streamlined—does not require the use of PCR primers that contain restriction sites

The pCR™2.1 vector provides:
• 3'-T overhangs for direct ligation of Taq-amplified PCR products
• T7 promoter for in vitro RNA transcription and sequencing
• A versatile polylinker with flanking EcoR I sites for easy excision of inserts
• M13 forward and reverse primer sites for sequencing

How TA Cloning® Works
Taq polymerase has a non-template-dependent activity that adds a single deoxyadenosine (A) to the 3' ends of PCR products. The linearized vector supplied in this kit has single 3' deoxythymidine (T) residues. This allows PCR inserts to ligate efficiently with the vector.

Kit Configurations
The TA Cloning® Kit is offered in a variety of configurations: without competent cells (K2020-20 and K2020-40), with One Shot® INVF' Chemically Competent E. coli (K2000-01 and K2000-40), with One Shot® TOP10F' Chemically Competent E. coli (K2030-01 and K2030-40), and with One Shot® TOP10 Chemically Competent E. coli (K2040-01 and K2040-40) in 20- and 40-reaction kit sizes.

GeneArt™ CRISPR Nuclease Vector with CD4 Enrichment Kit (with competent cells) (Invitrogen™)

The GeneArt® CRISPR Nuclease Vector with CD4 Enrichment Kit is a vector system for expression of the functional components needed for CRISPR/Cas9 genome editing in mammalian cells with a CD4 reporter. The CD4 reporter enables bead-based enrichment, an option for magnetic bead-based sorting/enrichment of Cas9 & CRISPR expressing cells using Dynabeads® CD4 magnetic beads. Transfection efficiency can also be tracked using anti-CD4 fluorescent antibodies. The linearized GeneArt® CRISPR nuclease vectors provide a rapid and efficient way to clone double-stranded oligonucleotides encoding a crRNA representing a desired target into an expression cassette that allows sequence-specific targeting of the Cas9 nuclease. This version of the kit includes OneShot® TOP10 Competent Cells. A version without competent cells is also available (Cat. No. A21175).

The GeneArt® CRISPR Nuclease Vector system with competent cells offers:

• Easy-to-design genome engineering system
• Affordable, ready-to-use cloning vectors
• Enrichment for hard to transfect or difficult to modify cell lines
• Competent cells included for added convenience and highest cloning efficiencies

All-in-One Vector System for CRISPR-based Genome Editing
The GeneArt® CRISPR Nuclease Vector kit offers a simple, ready-to-use, all-in-one expression vector system consisting of both a Cas9 nuclease expression cassette and a guide RNA (gRNA) cloning cassette for rapid and efficient cloning of DNA that encodes target-specific CRISPR RNA (crRNA). This system allows you to edit and engineer a genomic locus of choice in a sequence-specific manner from a single plasmid. After relevant targets have been identified with fast and easy-to-use GeneArt® CRISPR vectors, the biologically relevant mutations can be precisely created with GeneArt® Precision TALs, with high specificity and low off-target effects.

Need assistance with CRISPR gRNA design?
Our new CRISPR Search & Design tool allows you to search our database of >600,000 predesigned CRISPR gRNAs in human and mouse genes or analyze your sequence of interest for de novo gRNA designs using our proprietary algorithms. CRISPR sequences are optimized for gene knockout and target the first three transcribed exons for each gene. Search results include the top 6 CRISPR sequences with PAM sites, exon maps with gRNA binding sites, and potential off-target binding sites for each CRISPR sequence. The tool will design the correct gRNA format for your preferred CRISPR-Cas9 product, including oligos for your GeneArt® CRISPR Nuclease Vector. Start designing today >

Resources
Demonstrated protocol: Engineering stem cells with CRISPR-Cas9
Demonstrated protocol: CRISPR-Cas9 microinjection in mice and zebrafish embryos

Gateway™ pENTR™ 3C Dual Selection Vector (Invitrogen™)

Gateway® entry vectors are designed to clone DNA sequences using restriction endonucleases and ligase to create a Gateway® entry clone. The resulting entry clone is ready for recombination with a destination vector to create an expression clone. New: pENTR™ Dual Selection vectors!

The Gateway® entry vectors (Table 1) offer the following:
• attL1 and attL2 sites for site-specific recombination of the entry clone with a Gateway® destination vector to ensure cloning of the gene of interest in the correct orientation for expression
• Kozak consensus sequence for efficient translation initiation in eukaryotic systems
• Ribosome binding site for efficient translation initiation in prokaryotic systems (pENTR™ 1A Dual Selection, pENTR™3C Dual Selection, and pENTR™11 Dual Selection vectors only)
• rrnB transcription termination sequences to prevent basal expression of the PCR product of interest in E. coli
• pUC origin for high-copy replication and maintenance of the plasmid in E. coli
• Kanamycin resistance gene for selection in E. coli
• The ccdB⁄chloramphenicol fusion gene located between the two attL sites for
o negative selection and
o Chloramphenicol selection in E. coli
• Kanamycin resistance gene for selection in E. coli

pCRE Tluc16-DD Vector for Luciferase Assays (Thermo Scientific™)

The pCRE Tluc16-DD Vector is a transcriptional reporter vector designed to monitor the activation of cAMP-binding protein (CREB) and cAMP-mediated signal transduction pathways in mammalian cells. The vector encodes the smallest known luciferase, TurboLuc™16 (Tluc16) luciferase, as the reporter under the control of a combination of an optimized minimal core promoter and 5 tandem repeats of the c-AMP response element (CRE).

• Intracellular Tluc16 luciferase gene, optimized for high expression in mammalian systems
• Dual-destabilization (DD) technology reduces accumulation of Tluc16 luciferase mRNA and protein in cells, enhancing the responsiveness of the assay
• Monitor the activation of cAMP-binding protein (CREB) and cAMP-mediated signal transduction pathways in mammalian cells

Tluc16 luciferase is a 16 kDa, novel, intracellular luciferase derived from the marine copedod Metridia luciferase family. The wild-type luciferase has been modified to reduce its size, increase its brightness, and enable its intracellular expression.

The pCRE Tluc16-DD luciferase expression vector also contains dual-destabilization (DD) technology that reduces accumulation of the Tluc16 luciferase mRNA and protein in cells, enhancing the responsiveness of the assay. A synthetic polyA terminator and a Transcriptional Pause Site (TPS) are included upstream of the cAMP response elements to minimize non-specific transcriptional read-through. The Tluc16 luciferase activity can be measured using the Thermo Scientific TurboLuc™ One-Step Glow Assay Kit.

T-REx™ Complete Kit, with pcDNA™4/TO© Vector (Invitrogen™)

A Tetracycline-Regulated Expression System without Viral Transactivators

The T-REx™ System yields higher levels of induced expression than any other regulated mammalian expression system. It utilizes the complete CMV promoter and adds control elements from the bacterial tetracycline resistance operon to effectively repress and derepress transcription from one of the strongest mammalian promoter sequences known (1,2).

Specific Activation
The T-REx™ System uses a repressor mechanism that blocks transcription from the powerful CMV promoter in the absence of tetracycline. Because the T-REx™ System elements do not use viral transactivators, you can achieve high-level expression from the complete CMV promoter without secondary, non-specific activation of host genes.

The T-REx™ Mechanism
The T-REx™ transcriptional control elements are provided through two tetracycline operator sequences (TetO2) that have been inserted between the TATA box of the CMV promoter and the transcriptional start site. The TetO2 sequence itself has no effect on expression. When the tetracycline repressor protein (TR) is present, it effectively binds the TetO2 sites and blocks transcription initiation. Tetracycline added to the culture medium binds to, and changes the conformation of, the TR protein. This change causes the TR protein to release the TetO2 sites, derepressing transcription from the CMV promoter. The result is high-level expression of the gene of interest. Expression levels can be modulated based on the tetracycline concentration and can be induced to levels that are achieved with constitutive CMV expression vectors. T-REx™ is a powerful inducible mammalian expression system that allows you to regulate expression from the complete human cytomegalovirus (CMV) enhancer-promoter.

T-REx™ inducible expression vectors offer the following features:

• Complete CMV enhancer-promoter sequence containing two copies of the tetracycline operator TetO2 sequence for high-level regulated expression
• Zeocin™ or hygromycin resistance gene for effective selection of stable mammalian cell lines
• Large multiple cloning site to simplify cloning

In addition, pcDNA™4/TO/myc-His offers a c-myc epitope for rapid detection of the recombinant protein with an Anti-myc Antibody and a polyhistidine (6xHis) sequence for simple purification of the recombinant protein with nickel-chelating resin and detection with Anti- His(C-term) Antibody.

The regulatory vector, pcDNA™6/TR, is provided for high-level expression of the tetracycline repressor (TR) protein. This vector expresses the Blasticidin resistance gene for rapid selection of mammalian cell lines that stably express the TR protein.

pcDNA™3.2-DEST Mammalian Expression Vector (Invitrogen™)

The pcDNA™ vectors are designed for high-level, constitutive expression in a variety of mammalian cell lines. The pcDNA3.2/V5-DEST vector offers the following key features:

•Cytomegalovirus (CMV) promoter for high-level expression
attR sites for Gateway® cloning, enabling recombination with attL-flanked fragments
•C-terminal V5 tag for easy detection
•Neomycin resistance gene for stable selection
•Ampicillin resistance gene and pUC origin for selection and maintenance in E. coli

Gateway® Cloning
To fit all of your expression needs, Invitrogen offers state-of-the-art Gateway® destination vectors for expression in E. coli, insect, yeast, or mammalian cells, as well as for production of native protein or N- or C-terminal fusion proteins. All Gateway® destination vectors have attR sites for recombination with any attL-flanked fragment, regardless of whether it is an entry clone or an Ultimate™ ORF Clone. The following table lists a variety of available destination vectors.

Additional materials required, available separately: Gateway® entry clone, appropriate Gateway® LR Clonase® enzyme mix, and reaction buffer.

pT7CFE1-NHis-GST-CHA Vector for Mammalian Cell-Free Protein Expression (Thermo Scientific™)

Thermo Scientific pT7CFE1-NHis-GST-CHA is a cloning plasmid optimized to use with the Thermo Scientific 1-Step Human In Vitro Protein Expression System for in vitro translation (IVT) of tagged fusion proteins. pT7CFE1-NHis-GST-CHA Vector is available with tandem affinity tags, 9xHis and GST at the N-terminus and HA tag at the C-terminus, to facilitate protein purification and detection. pT7CFE1-NHis-GST-CHA also has a cleavable tag, HRV 3C, available on the N-terminus.

Features of pT7CFE1:
• EMCV IRES at the 5' UTR promotes high-level translation of mRNAs
• MCS accommodates gene insertion via ten different restriction sites: Msc1, Nde1, BamH1, EcoR1, EcoRV, Pac1, Pst1, Sac1, Sal1, Not1 and Xho1
• Poly A sequence in the 3' region promotes mRNA stabilization and protection from nucleases
• T7 terminator ensures synthesis of accurate size mRNA transcripts
• Plasmid linearization may be accomplished with restriction sites between Poly A sequence and the T7 terminator region

pT7CFE1 Expression Vectors contain the Encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) element that is critical for high levels of cap-independent protein expression in the Human In Vitro Translation System. Each vector features a highly-compatible multiple cloning site (MCS) to facilitate easy insertion of protein coding sequences into and between vectors. The pT7CFE1 Vector is available with single or tandem affinity tags at the N- or C- terminus to facilitate protein purification and detection. The pT7CFE Vectors are suitable for insertion of cloned genes, cDNAs, ORFs or PCR products for in vitro transcription and translation. Custom cloning services are also available.

More Product Data
Choosing a vector and purification method for in vitro protein expression

Related Products
pT7CFE1-NHA-CHis Vector for Mammalian Cell-Free Protein Expression
pT7CFE1-CGST-HA-His Vector for Mammalian Cell-Free Protein Expression
pT7CFE1-CGFP-HA-His Vector for Mammalian Cell-Free Protein Expression
pT7CFE1-NHis-GST Vector for Mammalian Cell-Free Protein Expression

Vivid Colors™ pLenti6.3/V5-GW/EmGFP Expression Control Vector (Invitrogen™)

The Vivid Colors™ pLenti6.3⁄V5-GW⁄EmGFP Expression Control Vector is a ViraPower™ HiPerform™ positive control lentiviral vector containing Emerald Green Fluorescent Protein (EmGFP). It is designed for use with the ViraPower™ HiPerform™ Lentiviral Expression Systems as a positive control to enable the detection of higher levels of EmGFP fluorescence following transfection in 293FT cells, as a titer control to produce an EmGFP-expressing lentivirus stock and as a transduction control following transduction in both dividing and non-dividing mammalian cells. The vector has the CMV promoter for driving constitutive expression of EmGFP and the SV40 promoter for driving expression of the blasticidin stable selection marker. The vector is equipped with two key genetic elements, making them Hiperform™ vectors: the Woodchuck Posttranscriptional Regulatory Element (WPRE) and the central Polypurine Tract (cPPT) sequence from the HIV-1 integrase gene to produce at least 4-fold increase in EmGFP expression compared to vectors lacking these elements. This control vector is not designed for generating EmGFP fusion proteins and does not express the V5 epitope.

Advantages
• Lentivirus based expression of EmGFP in dividing and non-dividing mammalian cells
• Serve as a quick positive control for transfection and lentiviral production
• Serve as a quick titer control in determination of lentivirus titer

Key Features
• Constitutive expression with CMV promoter
• High level expression of EmGFP without V5 epitope
• WPRE and cPPT sequences produce at least 4-fold increase in EmGFP expression compared to other Lenti vectors without these elements
• Blasticidin selection marker for stable selection

Kit Includes
• pLenti6.3⁄V5-GW⁄EmGFP Expression Control Vector

Related SKUs
• ViraPower™ HiPerform™ Lentiviral TOPO® Expression Kit (K531000)
• ViraPower™ HiPerform™ Lentiviral Gateway® Expression Kit (K533000)
• ViraPower™ HiPerform™ T-Rex™ Gateway® Expression System (A11141)
• ViraPower™ HiPerform™ Promoterless Gateway® Expression System (A11145)

For research use only. Not intended for any therapeutic or diagnostic use.

pFRT/lacZeo Vector (Invitrogen™)

pFRT/lacZeo and pFRT/lacZeo2 were constructed for establishing Flp-In™ Cell Lines that contain a single FRT site. Each vector includes a:

• FRT site for subsequent targeted integration of a Flp-In™ expression vector
Z-Zeocin™ fusion for selection of stable integrants with Zeocin™

These vectors differ in the SV40 promoter that drives expression of the lacZ-Zeocin™ fusion. pFRT/lacZeo2 contains a truncated version of the SV40 promoter. Use of this vector facilitates the isolation of clones that have integrated the vector near enhancer elements in the genome. Selection of stable clones using pFRT/lacZeo2 may allow easier isolation of clones that express a gene of interest at higher levels.

pTracer™-CMV/Bsd Mammalian Expression Vector (Invitrogen™)

Two Tracer™ mammalian expression vectors are available that express GFP fused to the selectable marker Blasticidin. Blasticidin is a potent selection agent that allows a more rapid generation of stable cell lines than other selection agents. Each vector uses a different set of promoters to express the gene of interest and the Cycle 3 GFP-Blasticidin resistance gene fusion.

The pTracer™-CMV/Bsd vector offers:

• CMV promoter for high-level constitutive expression of your gene of interest
• EF-1 promoter for high-level constitutive expression of the Cycle 3 GFP-Blasticidin resistance fusion

The pTracer™-EF/Bsd vectors offer:
• EF-1 promoter for high-level constitutive expression of your gene of interest
• CMV promoter for high-level constitutive expression of the Cycle 3 GFP-Blasticidin resistance fusion
• C-terminal V5 epitope tag for simple detection of recombinant protein with an Anti-V5 Antibody
• C-terminal 6xHis tag for rapid purification on nickel-chelating resin and simple detection with an Anti-His(C-term) Antibody
• Three reading frames for easier in-frame cloning

pJTI™ R4 Dest CMV N-EmGFP pA Vector

The pJTI™ R4 Dest CMV N-EmGFP pA vector allows a gene of interest to be expressed as an N-terminal Emerald Green Fluorescent Protein (EmGFP) fusion in a Jump-In™ parental cell line. When used in combination with a Jump-In™ parental cell line kit such as the Jump-In™ GripTite™ HEK293 cell line (A14150), an isogeneic stable cell line can be created with less effort and in less time than traditional cell engineering methods. In addition, Gateway® technology increases the efficiency of the cloning steps to introduce your GOI into the Gateway®-ready pJTI™ R4 Dest CMV N-EmGFP pA vector. The high retargeting efficiency, made possible by the R4 sites in the Jump-In™ parental 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 easy selection of a positive stable clone for expressing your gene of interest with an N-terminal EmGFP.

The Jump-In™ Vectors and Parental Cell Lines Let 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 due to the simplified work flow.
• Easily access the Jump-In™ technology with the ability to generate an unlimited number of cell lines without complicated licenses or restrictions to interpret.

Save Time with Rapid and Efficient Generation of Engineered Cell Lines
Using Gateway® technology, you can generate an expression construct for retargeting using the pJTI R4 DEST CMV N-EmGFP pA destination vector. In combination with a Jump-In™ parental cell line kit, the expression cassette is efficiently and specifically inserted into a genomic target R4 site resulting in the generation of functional cell pools in as little as 2 weeks without laborious clone isolation and analysis. Even generation of clonal cell lines can be done with reduced time and effort due to the high percentage of positive clones.

Expand Your Experimental Capabilities
The pJTI™ R4 Dest CMV N-EmGFP pA vector along with a Jump-In™ parental cell line provide the ideal solution for cells and assays where transient engineering technologies are problematic, as well as for difficult to "engineer" cell lines. 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.

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

Zero Blunt™ TOPO™ PCR Cloning Kit, with pCR™-Blunt II-TOPO™ Vector, One Shot™ TOP10 Chemically Competent E. coli, and PureLink™ Quick Plasmid Miniprep Kit (Invitrogen™)

Zero Blunt® TOPO® PCR Cloning Kits offers the fastest and easiest method for high-efficiency (cloning of blunt-end PCR products amplified with proofreading thermostable polymerases. The kits include linearized and topoisomerase I-activated pCR™-Blunt II-TOPO® Vector for 5-minute benchtop ligations without ligase (1). The pCR™-Blunt II-TOPO® Vector also features:

ccdB gene for positive selection (2)
EcoR I sites flanking the PCR product insertion site for easy excision of inserts
• Kanamycin and Zeocin™ resistance genes for your choice of selection in E. coli
• SP6 promoter/primer site for in vitro RNA transcription and sequencing
• M13 forward and reverse primer sites for sequencing or PCR screening

Selected Zero Blunt® TOPO® PCR Cloning Kits are available in combo kits combined with a PureLink™ Quick Plasmid Miniprep Kit (50 preps) for fast plasmid purification of your TOPO®-cloned inserts for downstream analysis.