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pTracer-EF/V5-His A A Vector Data

pTracer-EF/V5-His C C Vector Data

pTracer-EF/V5-His B B Vector Data

Agrin mediates cell contact-induced acetylcholine receptor clustering. Citations & References

  • Authors: Campanelli JT, Hoch W, Rupp F, Kreiner T, Scheller RH
  • Journal: Cell (1991) 67:909-916
  • PubMed ID: 1659950
Catalog #

pcDNA3.1D/V5-His-TOPO Vector Data

pcDNA3.1D/V5-His /LacZ Vector Data

Functional domains and DNA-binding sequences of RFLAT-1/KLF13, a Krüppel-like transcription factor of activated T lymphocytes. Citations & References

  • Authors: Sakamoto K, Yamaguchi S, Ando R, Miyawaki A, Kabasawa Y, Takagi M, Li CL, Perbal B, Katsube K.
  • Journal: J Biol Chem (2002) 277:30055-30120
  • PubMed ID: 12050170

alpha-Synuclein protects against oxidative stress via inactivation of the c-Jun N-terminal kinase stress-signaling pathway in neuronal cells. Citations & References

  • Authors: Hashimoto Makoto; Hsu Leigh J; Rockenstein Edward; Takenouchi Takato; Mallory Margaret; Masliah Eliezer;
  • Journal: J Biol Chem (2002) 277:11465-11537
  • PubMed ID: 11790792
Catalog #

Cloning and expression of a novel Na(+)-dependent neutral amino acid transporter structurally related to mammalian Na+/glutamate cotransporters. Citations & References

  • Authors: Shafqat S, Tamarappoo BK, Kilberg MS, Puranam RS, McNamara JO, Guadano-Ferraz A, Fremeau RT Jr
  • Journal: J Biol Chem (1993) 268:15351-15355
  • PubMed ID: 8340364
Catalog # K206001

Vivid Colors pcDNA6.2/EmGFP-Bsd/V5-DEST Mammalian Expression Vector Kit Manual / Product Insert

  • Version: Version C 29 October 2010
Catalog # V36620

Do any of the your mammalian expression vectors contain an SV40 intron? Product FAQ

Answer

We used to sell pcDNA1 vector and related vectors which contained the SV40 intron and polyA signal fragment downstream of the multiple cloning site. However, the SV40 intron shows "promiscuity", which means that in vectors containing the SV40 intron downstream of an expressed gene, the 3' end of the SV40 intron tends to splice with sequences within genes cloned upstream of the intron that are near-consensus 5' splice sites (consensus sequence is MAGGTRAGT), leading to aberrant splice products. In other words, the 3' end of the intron would often combine and splice out part of the gene of interest (1).

Our expression vectors now use the Bovine Growth Hormone polyadenylation sequence as the polyadenylation sequence for cloned genes of interest, because it is extremely efficient (2). However, the SV40 polyA signal is still included in many of our vectors as a polyadenylation signal for the expressed eukaryotic selection marker (e.g. neomycin, hygromycin, or zeocin resistance genes). There is no intron accompanying the polyA in these vectors.

(1) Huang MT, Gordon CM. The simian virus 40 small-t intron, present in many common expression vectors, leads to aberrant splicing.Mol Cell Biol. 1990 Apr;10(4):1805-10. PMID: 1690852).
(2) Goodwin, EC, Rottman, FM. The 3'-flanking sequence of the bovine growth hormone gene contains novel elements required for efficient and accurate polyadenylation.J Biol Chem. 1992 Aug 15;267(23):16330-4. PMID: 1644817

Answer Id:: E3952

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I am interested in a mammalian expression system where I can have regulated expression of my gene of interest. I see that you offer multiple systems for this purpose. Can you describe the main features of each system? Product FAQ

Answer

We offer three unique mammalian expression systems for inducible/regulated expression of the gene of interest:

- T-REx system
- Flp-In T-REx system
- GeneSwitch system

Please see below to see how they compare with one another:
System -- Basal Expression Level -- Induced Expression Level -- Response time to Maximal Expression -- Transgenic Appliation
T-Rex system -- Low -- Highest -- High -- Suitable
Flp-In T-REx system -- Lower -- High -- 24-48 hrs -- Suitable
GeneSwitch system -- Lowest -- High -- 24-48 hrs -- Suitable

Answer Id:: E9159

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Does the orientation of the SV40 polyadenylation region in your mammalian expression vectors affect expression? Product FAQ

Answer

The SV40 polyA is a region of the SV40 genome where transcripts coming from both directions terminate; both Small and Large T antigens from the counterclockwise direction and VP1, VP2 and VP3 from the clockwise direction. The fragment functions as a transcription terminator and poly A signal in either orientation.

Answer Id:: E3439

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I am cloning my gene of interest into the pENTR/SD/D-TOPO vector, but plan on expressing my gene in both E.coli and mammalian cells. Will the Shine-Dalgarno sequence interfere with mammalian expression? Product FAQ

Answer

No, the Shine-Dalgarno sequence does not adversely affect mammalian expression when used in an appropriate mammalian DEST vector.

Answer Id:: E6815

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I transduced my lentiviral stock into my mammalian cell line but am getting no expression of my gene of interest. What could have gone wrong? Product FAQ

Answer

Here are some possible causes and solutions:
- Promoter silencing; CMV promoter is prone to silencing especially in mouse and rat cells, screen multiple antibiotic resistant clones and select the one with the highest expression levels
- Viral stocks stored incorrectly; aliquot and store at -80 degrees C, do not freeze/thaw more than 3 times

Answer Id:: E9341

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