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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

pAd/BLOCK-iT™-DEST RNAi Gateway 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.

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.

pAd/CMV/V5-DEST™ Gateway™ Vector Kit (Invitrogen™)

The pAd⁄CMV⁄V5-DEST™ Gateway® Vector Kit contains the Gateway®-adapted ViraPower™ adenoviral expression vector, pAd⁄CMV⁄V5-DEST™ vector for easy recombination-based cloning and adenoviral-based, transient expression of a target gene in dividing and non-dividing mammalian cells. The vector allows generation of an adenovirus containing the target gene where constitutive, high-level expression is driven by the CMV promoter.

Advantages
• High efficiency and rapid recombination cloning
• Produces high titer adenoviral stocks
• Efficient delivery of the gene to dividing and non-dividing mammalian cells in vitro or in vivo
• Produces replication-incompetent virus for enhanced biosafety of the system
• Amenable for use in high-throughput applications

Key Features
• Gateway® Technology for efficient and rapid cloning
• CMV promoter for high-level constitutive expression of gene of interest
• Human Ad5 sequences (ΔE3) and Viral Inverted Terminal Repeats (ITRs) for packaging of the expression construct into virions
• Herpes Simplex Virus thymidine kinase (TK) polyadenylation sequence for efficient transcription termination and polyadenylation of mRNA
• V5 epitope for detection of recombinant protein
• Ampicillin selection marker

Kit Includes
• pAd⁄CMV⁄V5-DEST™ Gateway® Vector
• pAd⁄CMV⁄V5-GW⁄lacZ control plasmid

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

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

pcDNA™6.2/V5-PL-DEST Mammalian Expression Vector (Invitrogen™)

The pcDNA™ vectors are designed for high-level, constitutive expression in a variety of mammalian cell lines. The pcDNA6.2/V5-pL-DEST vector offers the following key features:

•Promoterless version of the pcDNA™6.2⁄V5-DEST vector (cat. no. 12489027)
attR sites for Gateway® cloning
•Compatible with MultiSite Gateway® Pro kits (e.g. cat. no. 12537100)
•C-terminal V5 tag for easy detection
•Blasticidin resistance gene for efficient 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.

pAd/PL-DEST™ Gateway™ Vector Kit (Invitrogen™)

The pAd⁄PL-DEST™ Gateway® Vector Kit contains the Gateway®-adapted ViraPower™ adenoviral expression vector, pAd⁄PL-DEST® vector for easy recombination-based cloning and adenoviral-based, transient expression of a target gene in dividing and non-dividing mammalian cells. The vector allows generation of an adenovirus containing the target gene where expression is driven by a promoter of choice. Alternatively, the vector may also be used to express small RNA molecules from their appropriate promoters.

Advantages
• High efficiency and rapid recombination cloning
• Produces high titer adenoviral stocks
• Efficient delivery of the gene to dividing and non-dividing mammalian cells in vitro or in vivo
• Allows gene of interest to be controlled by a promoter of choice
• Produces replication-incompetent virus for enhanced biosafety of the system
• Amenable for use in high-throughput applications

Key Features
• Gateway® Technology for efficient and rapid cloning
• Promoterless vector that allows gene of interest to be controlled by a promoter of choice
• Human Ad5 sequences (ΔE3) and Viral Inverted Terminal Repeats (ITRs) for packaging of the expression construct into virions
• Ampicillin selection marker

Kit includes
• pAd⁄PL-DEST™ Vector
• pAd⁄CMV⁄V5-GW⁄lacZ control plasmid

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

pCEP4 Mammalian Expression Vector (Invitrogen™)

These vectors are designed for high-level, constitutive expression from either the CMV or RSV promoters. Both vectors contain the EBNA-1 gene for episomal expression in primate and canine cell lines.

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.

Epi5™ Episomal iPSC Reprogramming Kit (Invitrogen™)

The Epi5™ Episomal iPSC Reprogramming Kit provides an easy-to-use, highly efficient set of 5 episomal vectors designed by Dr. Okita in the laboratory of Professor Yamanaka at the Center for iPS Cell Research and Application (CiRA), Kyoto University. This system produces transgene-free, virus-free human induced pluripotent stem cells (iPSCs) with efficiencies in the range of 0.04% to 0.3%, depending upon the cell type being reprogrammed.

Features of the Epi5™ Episomal iPSC Reprogramming Kit include:

• Easy to use and highly efficient—no need for small molecules during reprogramming and the added p53 suppression provides enhanced iPSC generation (1)
• Usable for all stages of your iPSC research—transgene-free and viral-free reprogramming allows use from basic to pre-clinical research
• Flexibility in media systems—can be used in either feeder-free or feeder-based media systems

Create Transgene and Virus-free iPSCs
Episomal vectors are a well-described system for producing transgene-free, virus-free iPSCs, providing a source of iPSCs for all stages of your pluripotent stem cell research. Other reprogramming methods, such as lentivirus, contain transgenes that can integrate into the host genome, potentially disrupting the genome or causing unpredictable results.

As oriP/EBNA1 vectors, these episomal vectors contain 5 reprogramming factors (Oct4, Sox2, Lin28, Klf4, and L-Myc) and replicate extra-chromosomally only once per cell cycle. At this replication rate, the episomes are lost at a rate of approximately 5% per cell generation. This system shows enhanced iPSC generation through p53 suppression, and the inclusion of L-Myc has been shown to be more potent and specific then c-Myc during human iPSC generation (1).

Easy-to-Use System
This 2-vial kit provides the optimal system for episomal vector reprogramming and eliminates the need for use of small molecules in the reprogramming medium. The episomal vectors are introduced into the cell by electroporation. We recommend use of the Neon®Transfection System for electroporation, allowing flexibility in the volume and thus number of cells needed for reprogramming.

Flexibility in Selection of Media Systems
The Epi5™ Episomal iPSC Reprogramming Kit can be used with multiple cell types and media systems. Protocols for fibroblasts and CD34+ blood cells are available in either feeder or feeder-free based reprogramming.

1. Okita K, Matsumara Y, Sato Y, et al. A more efficient method to generate integration free human iPS cells. Nature Methods 8, 409-412, 2011.

Zero Blunt™ PCR Cloning Kit (Invitrogen™)

The Zero Blunt® PCR Cloning Kit offers an easy method for high-efficiency (>80%) cloning of blunt-end PCR products amplified with proof-reading, thermostable DNA polymerases. The Zero Blunt® PCR Cloning Kit uses the multipurpose cloning vector pCR™-Blunt and ExpressLink™ T4 DNA Ligase to generate a ligation product in a five-minute, room-temperature ligation step.

Features of the Zero Blunt® Cloning® Kit with pCR™-Blunt vector:
Fast & convenient—5-minute, room-temperature ligation
EfficientccdB gene for positive selection results in >80% clones with correct insert
Flexible—choice of kanamycin or Zeocin™ resistance for flexible antibiotic selection

The pCR™-Blunt vector provides:
EcoR I sites flanking the PCR product insertion site for excision of inserts
• T7 promoter/primer site for in vitro RNA transcription and sequencing
• M13 forward and reverse primer sites for sequencing or PCR screening

How Zero Blunt® PCR Cloning Works
The Zero Blunt® PCR Cloning Kit is designed to clone blunt PCR fragments (or any blunt DNA fragment) with a low background of non-recombinants. The pCR™-Blunt vector contains the lethal E. coliccdB gene fused to the C-terminus of LacZα (Bernard et al., 1994). Ligation of a blunt PCR fragment disrupts expression of the lacZα-ccdB gene fusion permitting growth of only positive recombinants upon transformation. Cells that contain non-recombinant vector are killed when the transformation mixture is plated.

Kit Configurations
The Zero Blunt® PCR Cloning Kit is offered in a variety of configurations: with One Shot® TOPO10 Chemically Competent E. coli (K2700-20 and K2700-40) and without competent cells (K2750-20 and K2750-40) in 20- and 40- reaction kit sizes.

pLenti6/V5 Directional TOPO™ Cloning Kit (Invitrogen™)

The pLenti6⁄V5 Directional TOPO® Cloning Kit contains the TOPO®-adapted ViraPower™ lentiviral expression vector, pLenti6⁄V5-D-TOPO® for quick PCR-based cloning and high-level expression of a target gene in dividing and non-dividing mammalian cells. The vector has the CMV promoter for driving high-level, constitutive expression of the target gene and the blasticidin selection marker for stable selection in mammalian cells.

Advantages
• High efficiency and rapid cloning
• Constitutive gene expression in dividing and non-dividing mammalian cells in vitro or in vivo
• Produces replication-incompetent virus for enhanced biosafety of the system

Key Features
• Directional TOPO® Cloning site for rapid and efficient directional cloning of blunt-end PCR products
• Rouse Sarcoma Virus (RSV) enhancer⁄promoter for Tat-independent production of viral mRNA in the producer cell line
• Modified HIV-1 5’ and 3’ Long Terminal Repeats (LTR) for viral packaging and reverse transcription
• HIV-1 psi (ψ) packaging sequence for viral packaging
• HIV Rev response element (RRE) for Rev-dependent nuclear export of unspliced viral mRNA
• (CMV) immediate early promoter for high-level constitutive expression of the gene of interest in mammalian cells
• C-terminal V5 epitope for detection of the recombinant protein
• Blasticidin (bsd) resistance gene for selection in E. coli and mammalian cells
• Ampicillin resistance gene for selection in E. coli
• pUC origin for high-copy replication and maintenance of the plasmid in E. coli

Kit includes
pLenti6⁄V5-D- TOPO® Reagents
One Shot® Stbl3™ Chemically Competent E. coli

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

Champion™ pET300/NT-DEST and pET301/CT-DEST Gateway™ Vector Kit (Invitrogen™)

The Champion™ pET300/NT-DEST and pET301/CT-DEST Gateway® Vector Kit is designed for rapid cloning with a Gateway® entry clone and subsequent high-level prokaryotic expression controlled by 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 (pET300/NT-DEST and pET301/CT-DEST, respectively) for convenient purification and detection (Figure 1). The vector kit is ideal for structural biologists who desire no or minimal modifications to their protein of interest. To maximize expression, use with MagicMedia™ E. coli Expression Medium.



Contents and Storage:
The Champion™ pET300/NT-DEST and pET301/CT-DEST Gateway® Vector Kit includes 6 µg each of pET300/NT-DEST and pET301/CT-DEST vectors and 10 µg of control vector. Store at -20“C. Guaranteed stable for 6 months when properly stored.

pcDNA™4/TO Mammalian Expression 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 illustrated in Figure 1. Two tetracycline operator sequences (TetO2) 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 (Figure 2). 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™3.1 (+) 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 forward-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