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(Plasmid #126765)


Item Catalog # Description Quantity Price (USD)
Plasmid 126765 Standard format: Plasmid sent in bacteria as agar stab 1 $85

This material is available to academics and nonprofits only.


  • Vector backbone
    pX330-like (without U6-sgRNA coding sequence)
  • Total vector size (bp) 8056
  • Vector type
    Mammalian Expression, CRISPR

Growth in Bacteria

  • Bacterial Resistance(s)
    Ampicillin, 100 μg/mL
  • Growth Temperature
  • Growth Strain(s)
    NEB Stable
  • Copy number
    High Copy


  • Gene/Insert name
  • Alt name
  • Species
    S. pyogenes
  • Insert Size (bp)
  • Mutation
    M495V; Y515N; K526E; R661Q; amino acids 1005-1013 replaced with two glycine
  • Promoter CBh
  • Tags / Fusion Proteins
    • 3xFLAG (N terminal on insert)
    • NLS (N terminal on insert)
    • NLS (C terminal on insert)

Cloning Information

  • Cloning method Restriction Enzyme
  • 5′ cloning site MluI (not destroyed)
  • 3′ cloning site Pfl23II (destroyed during cloning)
  • 5′ sequencing primer GAACATCGTGATCGAAATG
  • 3′ sequencing primer TGTCGAAGCCGCCGTAC
  • (Common Sequencing Primers)

Terms and Licenses

  • Academic/Nonprofit Terms
  • Industry Terms
    • Not Available to Industry
  • Zeocin® is an InvivoGen trademark.

Depositor Comments

pCbh-3xFLAG-NLS-Blackjack-evoSpCas9-NLS (without U6-sgRNA coding sequence, with silent mutations).

Blackjack variants cleave both 20nt and 5'-extended 21nt spacer containing sgRNAs. B-evoSpCas9 shows higher fidelity and target selectivity than its parental increased fidelity variant, evoSpCas9.

The lack of sgRNA expression cassette allows for easy testing of various SpCas9 variants with the same sgRNA expressed from a separate plasmid (e.g. pmCherry_gRNA, Addgene# #80457).

For detailed information and plasmid usage, please see the associated publication.

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:

    B-evoSpCas9 was a gift from Ervin Welker (Addgene plasmid # 126765 ; ; RRID:Addgene_126765)
  • For your References section:

    Blackjack mutations improve the on-target activities of increased fidelity variants of SpCas9 with 5'G-extended sgRNAs. Kulcsar PI, Talas A, Toth E, Nyeste A, Ligeti Z, Welker Z, Welker E. Nat Commun. 2020 Mar 6;11(1):1223. doi: 10.1038/s41467-020-15021-5. 10.1038/s41467-020-15021-5 PubMed 32144253