pKK-FLAG-TEV
(Plasmid #105768)

• Purpose: (Empty Backbone) Expression of your protein of interest in fusion with FLAG at the N-terminus. The tag is cleavable by TEV protease or enterokinase.
• Depositing Lab: Andrzej Dziembowski
• Publication: Szczesny et al PLoS One. 2018 Mar 28;13(3):e0194887. doi: 10.1371/journal.pone.0194887. eCollection 2018.

Backbone

• Vector backbone: pcDNA5/FRT/TO
• Backbone manufacturer: Invitrogen
• Vector type: Mammalian Expression ; Flp-In competent
• Selectable markers: Hygromycin
• Tag / Fusion Protein:
  • FLAG-TEV (N terminal on backbone)

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: None
• Tag / Fusion Protein:
  • FLAG-TEV (N terminal on backbone)

Resource Information

• Supplemental Documents:
  • pKK-FLAG-TEV.txt
• Article Citing this Plasmid:
  • 1 Reference

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

This vector is a member of the pKK vector family. Its cloning site was optimized for the SLIC approach, however, a traditional restriction and ligation based approach is also possible (see Szczesny RJ et al. for detailed protocols; bioRxiv, doi: https://doi.org/10.1101/160101) This vector is a derivate of the pcDNA5/FRT/TO plasmid (Invitrogen) and is suitable for stable cell line generation using the Flp-In system. Transcription of the gene of interest is driven by a tetracycline responsive CMV promoter. A given coding sequence can be cloned into all pKK vectors using just three universal PCR primers and a ligation-independent DNA cloning method. FLAG is a short tag, less likely to interfere with protein function. FLAG tag present at the N-terminus of a protein is cleavable by enterokinase (PMID: 11694294). This is not the case if FLAG is present at the C-terminus of a protein.

Note: This plasmid contains an A211T mutation in AmpR and S84C mutation in HygR. The depositor has confirmed that this does not affect plasmid function.