This plasmid is designed to express tagged proteins in mammalian cells either by transient transfection or by creating stable cell lines. It contains a puromycin resistance expression cassette using the human Ubiquitin promoter to drive expression and allow for the selection of cells containing the plasmid.
About the Peptide Tag:This plasmid contains an Human Insulin (iSP) secretory signal peptide (SP) to allow proteins to be exported from the cytosol. During translocation from the cytosol the signal peptide is removed from the protein by endogenous proteases. This plasmid also contains a secondary Strep protein tag. The sequence of this tag is: WSHPQFEKWe provide a range of dual peptide tag plasmids. This is because some peptide tags provide specific biological properties (e. g., small molecule affinity new epitopes solubility or protein secretion) that are not provided by others.
About the Cleavage Tag: This plasmid also encodes a protease cleavage site that is designed to be positioned between your gene of interest and the tag to allow the removal of the tag following protein purification or isolation. This plasmid contains a TEV cleavage tag. The protein sequence of the cleavage tag is: ENLYFQG. Cleavage occurs between the Glu and Gly residues. TEV is often reported to have better specificity for its recognition site compared to EKT Thrombin or Faxtor Xa.
Promoter Expression Level: This plasmid contains the mammalian CMV promoter to drive gene expression. We have tested all of our mammalian promoters in a range of cell types and CMV is consistently the strongest in those we have studied. However there are many reports of the CMV promoter demonstrating silencing by methylation in long-term culture.
Looking for more vector options to move your experiments forward faster? Consider a custom cloning vector designed and built by Oxford Genetics™. Find out more at Oxford Genetics - Sigma's partner for cloning and expression vectors for molecular biology and synthetic biology applications.
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