The pUB6/V5-His Expression Vector
This description of the vector is purely for academic purposes.  If you are interested in purchasing this product
or learning about similar products, then click here to be linked to the Invitrogen web page.
 

    Molecular biologists utilize vectors to express and transport their genes of interest into bacterial cells.  A typical vector contains an origin of replication, a polylinker containing many restriction sites, and a reporter gene that translates proteins for antibiotic resistance.  These features permit the foreign DNA to be replicated and inserted, and the vector's presence in a bacterial cell to be observed.1
    Invitrogen has engineered a state-of-the-art vector, pUB6/V5-His.  The vector contains a promoter from the human ubiquitin C (UbC) gene which the company claims can express "recombinant proteins in a wider range of tissues in transgenic animals than other strong mammalian promoters". 2 The UbC promoter is just one feature among many that make it an excellent choice for the expression of recombinant proteins.

   
      In order to express recombinant fusion proteins, the gene must be placed in a reading frame with the C-terminal peptide.  The vector is produced in three different reading frames: A, B, and C.  As Figure 1 shows, the pUB6/V5-His vector contains many elements.  A description of each element follows.

    PUbc (human ubiquitin C promoter)-  This promoter allows recombinant proteins to be expressed in a wide range of mammalian cell lines.  Most vectors do not allow such broad levels of expression because of inconsistencies across species.  The ubiquitin C protein is highly conserved in eukaryotic cells; it can easily be introduced into a variety of cells.4
    V5 epitope-  This epitope is recognized by the Anti-V5 Antibody.  This epitope is translated as a part of the gene of interest.  Such epitope tags are helpful when discriminating between a wild type protein and a re-engineered protein of interest.  The epitope's amino acid sequence is:

-Gly-Lys-Pro-Ile-Pro-Asn-Pro-Leu-Leu-Gly-Leu-Asp-Ser-Thr-4
    C-terminal polyhistidine tag (6XHis)-  This epitope is recognized by the Anti-His(C-term) Antibody.  These six tandem histidine residues also aid in the purification of the recombinant proteins.5  To learn more about the Express system in which 6XHis is involved, click here.
    Bovine Growth Hormone and Polyadenylation signal (BGH pA)-  This signal is used during mRNA processing.  A polyadenine tail is added to the 3' end to protect the mRNA from hydrolytic enzymes and to provide stability.  It equally serves as a sequence for the termination of transcription.1  
    SV40 early promoter and origin (SV40 ori)-  Cells that are infected with the simian virus express the SV40 large T antigen.  This antigen prohibits the replication of the gene of interest.  The SV40 origin permits replication in such cells, assuring overexpression of the recombinant gene.3 
   EM-7 Promoter (EM-7)-  This promoter is necessary for the expression of the blasticidin resistance gene (see below).4
   Blasticidin Resistance Gene (BSD)-  This gene confers resistance to cells in which blasticidin is present.  Blasticidin is a nucleoside antibiotic that inhibits the formation of proteins in all cell types (prokaryotic and eukaryotic).  This feature is used to detect stable cell lines that are grown on blasticidin plates. 4
   SV40 polyadenylation signal (SV40 pA)- This signal is used during mRNA processing.  A polyadenine tail is added to the 3' end to protect the mRNA from hydrolytic enzymes and to provide stability.  It equally serves as a sequence for the termination of transcription.3
    pMB1 origin (pUC-derived) (pMB1 ori)-  This origin of replication promotes overexpression of the protein.
    Ampicillin Resistance Gene (Ampicillin)-  This is an alternative to the blasticidin resistance gene.  Cells are grown on ampicillin plates.  This feature is used to detect colonies containing the vector.1

    This combination of features creates a valuable tool for the expression of proteins.  The vectors that we have studied have not included such a variety of elements.  For example, we have mainly seen the ampicillin resistance gene.  This vector contains two: blasticidin and ampicillin.  By providing both of these resistance genes, the vector increases the accuracy of cell selection.  The vector provides several epitopes for the protein.  The most significant is the 6x-His tag (see above).  Above all, this vector allows protein expression across a wide range mammalian cell lines.  This element enhances our ability to look at gene conservation across species.  Using the same vector to look at the expression of one gene in several species might increase the proficiency of our research.
 
1.  Campbell, N.A.  1996.  Biology, Fourth Edition.  Menlo Park, CA:  Benjamin/Cummings Publishing Company
      p. 370-374.

2.  Expressions Newsletter, 1998.  Volume 5, Issue 6.  Carlsbad, CA:  Invitrogen Corporation. p. 15.

3.  "pTracer-SV40."  1998.  Invitrogen Web Resource - Invitrogen Web Resource - Manual.
    <http://www.invitrogen.com/pdf_manuals.html_ptracersv40_man.pdf> Accessed Feb 9, 1999.

4.  "pUB6/V5-His A, B, and C."  1998.  Catalog no.V250-20.   Invitrogen Web Resource - Manual.
    <http://www.invitrogen.com/pdf  manuals.html_pub6v5his_man.pdf>  Accessed Feb 9, 1999.

5.  "XpressTM."  1998.  Catalog no.R801-01.   Invitrogen Web Resource - Brochure.
    <http://www.invitrogen.com/manuals.html_xpress_bro.pdf>  Accessed Feb 9, 1999.

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