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TC-FlAsH™ TC-ReAsH™ II In-Cell Tetracysteine Tag Detection Kit, with Mammalian TC-Tag Gateway™ Expression Vectors (Green Fluorescence) (Red Fluorescence) (Invitrogen™)

TC-FlAsH™ TC-ReAsH™ II In-Cell Tetracysteine Tag Detection Kit is an expression tag-based fluorescence labeling system for live-cell labeling. The expression construct is created when the Gateway entry clone (containing the gene of interest) is recombined with one of two destination vectors (pcDNA6.2/nTC-Tag-DEST for N-terminal tagging or pcDNA6.2/cTC-Tag-DEST for C-terminal tagging). The expression construct is then used to transform the host strain. The protein can be detected either with green-fluorescent FlAsH-EDT2 reagent or red-fluorescent ReAsH-EDT2 reagent. The tag is very small (6 amino acids) and the protein of interest is fluorescent only when the labeling reagent is added. BAL wash buffer replaces the previously supplied Disperse Blue 3 and EDT-based wash buffer as an olfactorily more agreeable reagent that yields superior signal to noise.

The kit contains FlAsH-EDT2 labeling reagent, ReAsH-EDT2 labeling reagent, BAL wash buffer, pcDNA6.2/cTC-Tag-DEST, pcDNA6.2/nTC-Tag-DEST, and pcDNA6.2/nTC-Tag-p64 Control Plasmid. BAL wash buffer should be stored at 4°C, and the rest of the kit stored at -20°C, protected from light. The kit is stable for 6 months when stored correctly.

Expressway™Lumio™ Expression and Detection System, without vector (Invitrogen™)

The Expressway™ Lumio™ Expression and Detection System takes advantage of the Lumio™ recognition sequence, a small, six amino acid sequence (Cys-Cys-Pro-Gly-Cys-Cys). The Lumio™ detection reagent binds the recognition sequence with high specificity and affinity, resulting in a bright fluorescent signal for real-time protein production analysis and immediate in-gel protein detection. In addition, Expressway’s specialized E. coli lysate, derived from a slyD mutant, eliminates nonspecific binding of the Lumio™ Green Detection Reagent to the endogenous SlyD protein and provides optimal background for detection of recombinant proteins (Figure 1). The Lumio™ Green Detection Kit is included in the system for real-time detection of protein synthesis as well as easy in-gel protein detection. The Lumio™ vectors (Figures 2 and 3) also feature:

attR sites for efficient recombination with any attL-flanked Gateway® entry vector
N-terminal Lumio™ tag (pEXP3-DEST vector) with TEV cleavage site for efficient removal of the Lumio™ sequence following purification
C-terminal Lumio™ tag (pEXP4-DEST vector) for easy, and immediate in-gel detection
T7 promoter, ribosome binding site, and T7 terminator optimally spaced for cell-free protein expression

pYES2/NT A, B, & C Yeast Expression Vectors (Invitrogen™)

pYES2/NT and pYES2/CT are S. cerevisiae expression vectors derived from the parental pYES2 vector. Both vectors offer the URA3 gene for selection in yeast. pYES2/NT features an N-terminal Xpress™ epitope for detection with Invitrogen's Anti-Xpress™ Antibody and a polyhistidine (6xHis) tag for purification with nickel-chelating resin. pYES2/CT contains a C-terminal V5 epitope for detection and a polyhistidine (6xHis) tag for purification.

pEF1/His A, B, & C Mammalian Expression Vectors (Invitrogen™)

All pEF or pUB vectors contain a strong promoter for high-level expression in mammalian cells, a choice of selection marker for generating stable cell lines, and an epitope tag for easy detection with a monoclonal antibody and rapid purification on nickel-chelating resin. Each vector is available in three reading frames to simplify cloning in-frame with the fusion tag.

pcDNA™3.1/His A, B, & C Mammalian Expression Vectors (Invitrogen™)

All pcDNA™ vectors contain a strong promoter for high-level expression in mammalian cells, a choice of selection marker for generating stable cell lines, and an epitope tag for easy detection with a monoclonal antibody and rapid purification on nickel-chelating resin. Each vector is available in three reading frames to simplify cloning in-frame with the fusion tag.

pMT/V5-His A, B, & C Drosophila Expression Vectors (Invitrogen™)

The DES®-Inducible Kit provides the expression vector pMT/V5-His for expression of recombinant proteins. This vector uses the Drosophila metallothionein gene promoter that is induced in S2 cells upon addition of copper sulfate or cadmium chloride to the culture medium.

The pMT/V5-His vector offers the following features:

• Small size (3.5 kb) to improve DNA yields and increase subcloning efficiency
• C-terminal V5 epitope tag for rapid detection with Invitrogen's Anti-V5 Antibody
• C-terminal 6xHis tag for simple purification of recombinant fusion proteins using nickel-chelating resin

To facilitate cloning, a set of three vectors-A, B, and C-is provided. Each vector has the multiple cloning site in a different reading frame relative to the coding sequence of the C-terminal tag.

pUB6/V5-His A, B, & C Mammalian Expression Vectors (Invitrogen™)

All pEF or pUB vectors contain a strong promoter for high-level expression in mammalian cells, a choice of selection marker for generating stable cell lines, and an epitope tag for easy detection with a monoclonal antibody and rapid purification on nickel-chelating resin. Each vector is available in three reading frames to simplify cloning in-frame with the fusion tag.

pGAPZ A, B, & C Pichia pastoris Expression Vectors (Invitrogen™)

pGAPZ A, B, & C and pGAPZ A, B, & C are expression vectors designed for high-level, constitutive expression in Pichia pastoris. pGAPZ and pGAPZ were created by replacing the methanol-regulated AOX1 promoter with the constitutive, glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter in the backbone of the pPICZ vectors. Although the yield
of any protein constitutively expressed in the Pichia system is dependent on the toxicity of the protein to yeast, constitutive expression under the control of pGAPZ or pGAPZ vectors can sometimes produce greater yields than inducible expression (1).

pcDNA™4/myc-His A, B, & C Mammalian Expression Vectors (Invitrogen™)

All pcDNA™ vectors contain a strong promoter for high-level expression in mammalian cells, a choice of selection marker for generating stable cell lines, and an epitope tag for easy detection with a monoclonal antibody and rapid purification on nickel-chelating resin. Each vector is available in three reading frames to simplify cloning in-frame with the fusion tag.

pGAPZα A, B, & C Pichia pastoris Expression Vectors (Invitrogen™)

pGAPZ A, B, & C and pGAPZ A, B, & C are expression vectors designed for high-level, constitutive expression in Pichia pastoris. pGAPZ and pGAPZ were created by replacing the methanol-regulated AOX1 promoter with the constitutive, glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter in the backbone of the pPICZ vectors. Although the yield
of any protein constitutively expressed in the Pichia system is dependent on the toxicity of the protein to yeast, constitutive expression under the control of pGAPZ or pGAPZ vectors can sometimes produce greater yields than inducible expression (1).

pTracer™-EF A, B, & C Mammalian Expression Vectors (Invitrogen™)

Three pTracer™ mammalian expression vectors are available that express GFP fused to the selectable marker Zeocin™ . Each vector uses a different set of promoters to express the gene of interest and the Cycle 3 GFP-Zeocin™ fusion.

All three pTracer™ vectors have the following features:

• Cycle 3 GFP-Zeocin™ fusion for selection in mammalian cell lines
• BGH polyA signal and transcription termination sequences to enhance mRNA stability

The pTracer™-SV40 vector offers:
• SV40 promoter for high-level constitutive expression of your gene of interest
• CMV promoter for high-level constitutive expression of the Cycle 3 GFP-Zeocin™ fusion
• SV40 origin for episomal replication and simple vector rescue in cell lines expressing the SV40 large T antigen

The pTracer™-CMV2 vector offers:
• CMV promoter for high-level constitutive expression of your gene of interest
• EF-1α promoter for high-level constitutive expression of the Cycle 3 GFP-Zeocin™ fusion
• Ampicillin resistance gene for selection in E. coli

The pTracer™-EF vectors offer:
• EF-1α promoter for high-level constitutive expression of your gene of interest
• CMV promoter for high-level constitutive expression of the Cycle 3 GFP-Zeocin™ fusion
• C-terminal V5 epitope tag for simple detection of recombinant fusion proteins 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
• Multiple cloning site supplied in three reading frames to simplify cloning in frame with the C-terminal tag

pTrcHis A, B, & C Bacterial Expression Vectors (Invitrogen™)

Our pTrcHis A, B, & C vectors are designed to offer enhanced translation initiation and high-level expression in E. coli. These vectors feature:

• High-level regulated transcription from the trc promoter
• Enhanced translation efficiency of eukaryotic genes in E. coli
• The lacO operator and lacIq repressor gene for transcriptional regulation in any E. coli strain

This particular vector offers:

• N-terminal polyhistidine (6xHis) tag for rapid purification with nickel-chelating resin and detection with an anti-HisG antibody
• N-terminal Xpress™ epitope for easy detection with an anti-Xpress™ antibody
• Enterokinase cleavage site for removal of fusion tag

For C-terminal polyhistidine tag and c-myc epitope, please see our pTrcHis2 A, B, & C Vector.

pPIC6α A, B, & C Pichia Vectors (Invitrogen™)

pPIC6 A, B, & C are Pichia pastoris expression vectors designed for simple selection,
high-level expression, and rapid protein purification and detection. The pPIC6 A,
B, & C vectors are designed for high-level secreted expression. Both sets of vectors
have the following features:
Blasticidin resistance for direct selection of multi-copy integrants
Inducible AOX1 promoter for high-level expression in Pichia pastoris
C-terminal c-myc epitope for convenient detection with an Anti-myc Antibody
C-terminal polyhistidine (6xHis) tag for rapid purification with nickel-chelating
resin and detection with an Anti-His(C-term) Antibody
pPIC6 also features:
An É -factor secretion signal for efficient transport of proteins to the medium

pSecTag2/Hygro A, B, & C Mammalian Expression Vectors (Invitrogen™)

pSecTag2 and pSecTag2/Hygro are mammalian expression vectors designed for the secretion, purification, and detection of fusion proteins. Each vector has a large multiple cloning site in three reading frames to simplify cloning in frame with the N-terminal secretion signal. The vectors (Figure 1) offer the following features:

• Secretion signal from the V-J2-C region of the mouse Ig kappa-chain for efficient secretion of recombinant proteins (Figure 2)
• Cytomegalovirus (CMV) promoter for high-level constitutive expression
• C-terminal polyhistidine (6xHis) tag for rapid purification with nickel-chelating resin and detection with an Anti-His(C-term) Antibody
• C-terminal c-myc epitope for detection with an Anti-myc Antibody
• Bovine Growth Hormone (BGH) polyadenylation signal and transcription termination sequence to enhance mRNA stability
• SV40 origin for episomal replication and simple vector rescue in cell lines expressing the large T antigen (e.g., COS-1, COS-7)

The pSecTag2 vectors carry the Zeocin™ resistance gene for cost-effective selection in mammalian cells. Zeocin™ selection can also be used in E. coli.

The pSecTag2/Hygro vectors have the Hygromycin B resistance gene for selection of stable mammalian cell lines.

Episomal iPSC Reprogramming Vectors (Invitrogen™)

Episomal iPSC Reprogramming Vectors are a cost-effective mixture of three vectors designed to provide the optimal system for generating transgene-free and virus-free induced pluripotent stem cells (iPSCs) in a feeder-free environment. Originally developed by Junying Yu and James Thomson(1) and further optimized by Cellular Dynamics International, these Episomal iPSC Reprogramming Vectors have proven successful in reprogramming a number of different somatic cell types.
• Safe for all stages of your iPSC research—transgene-free and viral-free reprogramming allows use from basic to pre-clinical research
• Reprogram a variety of somatic cell types—provides flexibility in somatic cell selection
• Optimized for feeder-free reprogramming—allows for defined and feeder-free reprogramming when used with Essential 8™ Medium

Create Transgene- and Virus-free iPSCs
The Episomal iPSC Reprogramming 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. These episomal vectors are introduced into the cell by electroporation. As oriP/EBNA1 vectors, they contain all 6 reprogramming factors (Oct4, Sox2, Nanog, Lin28, Klf4 and lMyc) and replicate extrachromosomally only once per cell cycle. At this replication rate, the episomes are lost at a rate of approximately 5% per cell generation.

Generate iPSCs from a Wide Variety of Somatic Cell Types
iPSCs have been generated with episomal vectors from a range of somatic cells including fibroblasts, bone marrow mononuclear cells, PBMCs, lymphoblast B cells, and various disease-type fibroblasts and PBMCs. Each kit provides enough material for 6 reprogramming experiments.

Optimized for Feeder-free Reprogramming with Essential 8™ Medium
The Episomal iPSC Reprogramming Vectors were designed in the laboratories of James Thomson and Cellular Dynamics International for use with Essential 8™ Medium, thus providing an optimal environment for defined, feeder-free reprogramming.

Commercialized in Partnership with Cellular Dynamics International.

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

Reference:
1. Human Induced Pluripotent Stem Cells Free of Vector and Transgene Sequences; Yu J, Hu K, Smuga-Otto K, Tian S, Stewart R, Slukvin II, Thomson J. Science. 2009; 324:797-801.