C-Terminus pQE Vector Set

For high-level expression of C-terminally 6xHis-tagged proteins
  • C-terminal 6xHis tag ensures that only full-length protein is purified
  • pQE-16 vector for expression of poorly expressed proteins as DHFR fusions
  • pQE-16 vector for expression of short peptides as DHFR fusions

This set provides 3 vectors (pQE-16, pQE-60, and pQE-70) for expression of C-terminally 6xHis-tagged proteins and is recommended for open reading frames with “pause sites”, which can cause premature termination. pQE-60 and pQE-70 allow the original start codon of the coding fragment to replace the ATG in the pQE vector, preserving the authentic N-terminus of the protein. These two constructs are created by introducing an NcoI and a SphI restriction site, respectively, at the ATG codon of the insert by PCR or mutagenesis. pQE-16 allows expression of C-terminally 6xHis-tagged DHFR-fusion proteins. DHFR (dihydrofolate reductase) enhances antigenicity and stability, and is recommended for poorly expressed proteins or short peptides prone to proteolysis. Since DHFR itself displays little immunogenicity in mouse and rat, DHFR-fusion proteins are ideal for epitope screening.

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C-Terminus pQE Vector Set
25 µg each: pQE-16, pQE-60, pQE-70
 32903
$877.00
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The C-Terminus pQE Vector Set is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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Principle

QIAexpress pQE vectors combine a powerful phage T5 promoter (recognized by E. coli RNA polymerase) with a double lac operator repression module to provide tightly regulated, high-level expression of recombinant proteins in E. coli (see figure "QIAexpress pQE vector"). Protein synthesis is effectively blocked in the presence of high levels of lac repressor and the stability of cytotoxic constructs is enhanced. The pQE vectors enable placement of the 6xHis tag at either the N- or C-terminus of the recombinant protein.

 

Elements present in QIAexpress pQE vectors
ElementDescription
Optimized promoter/operator element Consists of the phage T5 promoter and two lac operator sequences, which increase the probability of lac repressor binding and ensure efficient repression of the powerful T5 promoter
Synthetic ribosomal binding site RBSII For efficient translation
6xHis-tag coding sequence Either 5' or 3' to the polylinker cloning region
Translational stop codons In all reading frames for convenient preparation of expression constructs
Two strong transcriptional terminators t0 from phage lambda, and T1 from rrnB operon of E. coli, to prevent read-through transcription and ensure stability of the expression construct
ColE1 origin of replication From pBR322
Beta-lactamase gene (bla) Confers ampicillin resistance

Procedure

Inserts encoding proteins of interest are cloned into appropriate constructs (for detailed information, see The QIAexpressionist Handbook) and transformed into a suitable E. coli strain for expression. Expression is induced by addition of IPTG. Vector pQE-TriSystem constructs can be transformed into E. coli, used as a shuttle vector for recombinant protein expression in insect cells, or transfected into mammalian cells.

Applications

The QIAexpress Expression system provides high-level expression of proteins suitable for
many applications, including:

  • Purification of functional, conformationally active proteins 
  • Purification under denaturing conditions for antibody production
  • Crystallization for determination of three-dimensional structure 
  • Assays involving protein–protein and protein–DNA interactions
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Kit Handbooks
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The QIAexpressionist - (EN)
A handbook for high-level expression and purification of 6xHis-tagged proteins
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