pTrcHis2 A, B, & C Bacterial Expression Vectors
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Citas y referencias (5)
Invitrogen™

pTrcHis2 A, B, & C Bacterial Expression Vectors

Nuestros vectores pTrcHis A, B y C están diseñados para ofrecer una iniciación de traducción mejorada y una expresión deMás información
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Número de catálogoCantidad
V3652020μg
Número de catálogo V36520
Precio (MXN)
-
Cantidad:
20μg
Nuestros vectores pTrcHis A, B y C están diseñados para ofrecer una iniciación de traducción mejorada y una expresión de alto nivel en E. coli. Estos vectores incluyen:

• transcripción regulada de alto nivel del promotor de trc
• Mejora de la eficacia de la traducción de genes eucarióticos en E. coli
• El operador de lacO y el gen represor de lacIq para la regulación transcripcional en cualquier cepa de E. coli

Este vector en particular ofrece:

• Etiqueta de polihistidina (6xHis) C-terminal para una rápida purificación con resina quelante de níquel y detección con anticuerpos anti-His (C-terminal)
• Epitopo c-myc C-terminal para una detección sencilla de proteínas de fusión con un anticuerpo myc

Para la etiqueta de polihistidina N-terminal y el epítopo Xpress™, consulte nuestro vector pTrcHis A, B y C.
Para uso exclusivo en investigación. No apto para uso en procedimientos diagnósticos.
Especificaciones
Resistencia bacteriana a los antibióticosAmpicilina (AMPR)
HendiduraSitio de reconocimiento de EK (enterocinasa)
Sistema constitutivo o inducibleInducible
Agente inductorIPTG
Tipo de productoVector de expresión bacteriana
Cantidad20μg
Agente de selección (eucariótico)Ninguno
VectorpTrc
Método de clonaciónEnzimas de restricción/MCS
PromotorTrc, lacO
Etiqueta de proteínaTiorredoxina
Unit Size20 µg
Contenido y almacenamiento
Todos los vectores son superenrollados y se suministran liofilizados. Se suministran una punción E. colistab TOP10 y un control de expresión positivo. Almacene los vectores a -20 °C. Almacenar las punciones a temperatura ambiente. Se garantiza la estabilidad de este vector durante seis meses si se almacena adecuadamente.

Preguntas frecuentes

What are the advantages and/or disadvantages of using TOP10, DH5α, or other cloning strains versus BL21 Star (DE3) or BL21 (DE3) cells for expression with the pTrc system?

Top10, DH5α, other cloning strains

Advantages:
- Saves time, can go directly from cloning to expression.
- The glycerol stock is more stable because these strains are endA- and recA-.

Disdvantages:
- If GOI is toxic, the cloning step can be difficult.
- These cloning strains are not protease-deficient; therefore, the protein may be degraded.

BL21 Star(DE3) or BL21 (DE3)

Advantages:
- These expression strains are protease-deficient.
- You have to transform the plasmid into an expression strain.
- The glycerol stock may be unstable because these expression strains are not endA- and recA-.
- The (DE3) part is wasted because the promoter does not need T7 RNA polymerase.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

What competent cell strain can I use for expression with my pTrc Expression System?

The pTrc promoter is recognized by E. coli RNA polymerase, not T7 polymerase, and therefore can be expressed in any E. coli strain, not just BL21 strains. Therefore, you can use Top10, DH5α, etc. for expression. However, if your gene is toxic, the cloning step can be difficult as there is leakiness in expression.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

How does glucose repress the pTrc promoter?

A transcriptional activator protein called CAP (catabolite activator protein) normally binds upstream of the trc promoter and activates transcription. This protein requires cAMP to bind the DNA. Adding glucose to the medium can reduce intracellular cAMP levels. Supplementing LB medium and agar plates with glucose will repress basal level transcription from the trc promoter. We recommend that you include 25 mM, 0.5% glucose in the selection medium to ensure stability of your insert.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

What are the benefits of the pTrc expression system?

The pTrc promoter in the pTrc system is a strong hybrid promoter composed of the -35 region of the trp promoter and the -10 region of the lacUV5 promoter/operator. Expression of pTrc is repressed by the LacI protein and induced by IPTG.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

Do I need to include a ribosomal binding site (RBS/Shine Dalgarno sequence) or Kozak sequence when I clone my gene of interest?

ATG is often sufficient for efficient translation initiation although it depends upon the gene of interest. The best advice is to keep the native start site found in the cDNA unless one knows that it is not functionally ideal. If concerned about expression, it is advisable to test two constructs, one with the native start site and the other with a Shine Dalgarno sequence/RBS or consensus Kozak sequence (ACCAUGG), as the case may be. In general, all expression vectors that have an N-terminal fusion will already have a RBS or initiation site for translation.

Find additional tips, troubleshooting help, and resources within our Protein Expression Support Center.

View all pTrcHis2 A, B, & C Bacterial Expression Vectors FAQs

Citations & References (5)

Citations & References
Abstract
Identification and characterization of galectin-9, a novel beta- galactoside-binding mammalian lectin.
Authors:Wada J, Kanwar YS
Journal:J Biol Chem
PubMed ID:9038233
'A 36-kDa beta-galactoside mammalian lectin protein, designated as galectin-9, was isolated from mouse embryonic kidney by using a degenerate primer polymerase chain reaction and cloning strategy. Its deduced amino acid sequence had the characteristic conserved sequence motif of galectins. Endogenous galectin-9, extracted from liver and thymus, as well as recombinant ... More
A role for RanBP1 in the release of CRM1 from the nuclear pore complex in a terminal step of nuclear export.
Authors:Kehlenbach RH, Dickmanns A, Kehlenbach A, Guan T, Gerace L
Journal:J Cell Biol
PubMed ID:10330396
'We recently developed an assay in which nuclear export of the shuttling transcription factor NFAT (nuclear factor of activated T cells) can be reconstituted in permeabilized cells with the GTPase Ran and the nuclear export receptor CRM1. We have now used this assay to identify another export factor. After preincubation ... More
HSP70-2 is required for CDC2 kinase activity in meiosis I of mouse spermatocytes [published erratum appears in Development 1997 Sep;134(17):3218]
Authors:Zhu D, Dix DJ, Eddy EM
Journal:Development
PubMed ID:9247342
'Cyclin B-dependent CDC2 kinase activity has a key role in triggering the G2/M-phase transition during the mitotic and meiotic cell cycles. The Hsp70-2 gene is expressed only in spermatogenic cells at a significant level. In Hsp70-2 gene knock-out (Hsp70-2(-/-)) mice, primary spermatocytes fail to complete meiosis I, suggesting a link ... More
Activation mechanism of Gi and Go by reactive oxygen species.
Authors: Nishida Motohiro; Schey Kevin L; Takagahara Shuichi; Kontani Kenji; Katada Toshiaki; Urano Yasuteru; Nagano Tetsuo; Nagao Taku; Kurose Hitoshi;
Journal:J Biol Chem
PubMed ID:11781308
Reactive oxygen species are proposed to work as intracellular mediators. One of their target proteins is the alpha subunit of heterotrimeric GTP-binding proteins (Galpha(i) and Galpha(o)), leading to activation. H(2)O(2) is one of the reactive oxygen species and activates purified Galpha(i2). However, the activation requires the presence of Fe(2+), suggesting ... More
Galectin-7 (PIG1) exhibits pro-apoptotic function through JNK activation and mitochondrial cytochrome c release.
Authors: Kuwabara Ichiro; Kuwabara Yasuko; Yang Ri-Yao; Schuler Martin; Green Douglas R; Zuraw Bruce L; Hsu Daniel K; Liu Fu-Tong;
Journal:J Biol Chem
PubMed ID:11706006
Galectin-7 is normally expressed in all types of stratified epithelia, but is significantly down-regulated in squamous cell carcinomas. This protein was recently found to be highly inducible by p53 in a colon carcinoma cell line, DLD-1, and designated as PIG1 (for p53-induced gene 1). We studied transfectants of HeLa and ... More