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pKK-BI16-ORF1-3C-mRuby3_ORF2-TEV-mClover3
(Plasmid #105802)

Ordering

Item Catalog # Description Quantity Price (USD)
Plasmid 105802 Standard format: Plasmid sent in bacteria as agar stab 1 $65

This material is available to academics and nonprofits only.

Backbone

  • Vector backbone
    BI-16 (constructed based on pcDNA5/FRT/TO from Invitrogen)
  • Backbone manufacturer
    Sammarco & Grabczyk (PMID: 16212928)
  • Vector type
    Mammalian Expression ; Flp-In competent
  • Selectable markers
    Hygromycin
  • Tag / Fusion Protein
    • ORF1: 3C-mRuby3; ORF2: TEV-mClover3 (C terminal on backbone)

Growth in Bacteria

  • Bacterial Resistance(s)
    Ampicillin
  • Growth Temperature
    37°C
  • Growth Strain(s)
    DH5alpha
  • Copy number
    High Copy

Gene/Insert

  • Gene/Insert name
    None
  • Tag / Fusion Protein
    • ORF1: 3C-mRuby3; ORF2: TEV-mClover3 (C terminal on backbone)

Resource Information

Depositor Comments

This vector is a member of the pKK-BI16 series which belongs to the pKK vector family. It enables concomitant expression of two genes, which transcription is driven by a tetracycline responsive bidirectional CMV promoter. This vector is a derivate of the BI16 plasmid (PMID: 16212928) and is suitable for stable cell line generation using the Flp-In system. ORF2 is cloned with the universal SLIC protocol (just three universal PCR primers are required to clone a given coding sequence into all vectors from pKK family by a ligation-independent DNA cloning method), ORF1 can be cloned using SLIC with specifically designed primers (applicable enzymes: Bsp120I, ApaI, MluI, NotI, BspTI, Eco47III, MunI) or conventional restriction-ligase based cloning (see Szczesny RJ et al. for detailed description and protocols; bioRxiv, doi: https://doi.org/10.1101/160101). mClover3 is a brighter derivative of mEGFP. mRuby3 is brighter than mCherry and has slightly shifted excitation and emission spectra. The mClover3-mRuby3 pair is superior to EGFP-mCherry in FRET analysis (PMID: 26879144). Useful for experiments requiring co-expression of two proteins (e.g. FRET measurements).

How to cite this plasmid ( Back to top)

These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the plasmids were described, and include Addgene in the Materials and Methods of your future publications.

  • For your Materials & Methods section:

    pKK-BI16-ORF1-3C-mRuby3_ORF2-TEV-mClover3 was a gift from Andrzej Dziembowski (Addgene plasmid # 105802 ; http://n2t.net/addgene:105802 ; RRID:Addgene_105802)
  • For your References section:

    Versatile approach for functional analysis of human proteins and efficient stable cell line generation using FLP-mediated recombination system. Szczesny RJ, Kowalska K, Klosowska-Kosicka K, Chlebowski A, Owczarek EP, Warkocki Z, Kulinski TM, Adamska D, Affek K, Jedroszkowiak A, Kotrys AV, Tomecki R, Krawczyk PS, Borowski LS, Dziembowski A. PLoS One. 2018 Mar 28;13(3):e0194887. doi: 10.1371/journal.pone.0194887. eCollection 2018. 10.1371/journal.pone.0194887 PubMed 29590189