pRubiC-T2A-Cas9
(Plasmid #75347)

• Purpose: Ubiquitin promoter expresses mCherry and humanized spCas9 (PX330) via a T2A motif. For cell transfection or use in retroviral (MMuLV) packaging. Use Pac1 and/or BstB1 sites to insert sgRNAs from PXL.
• Depositing Lab: Bryan Luikart
• Publication: Williams et al Sci Rep. 2016 May 10;6:25611. doi: 10.1038/srep25611.

Backbone

• Vector backbone: pRubi
• Backbone size w/o insert (bp): 6939
• Total vector size (bp): 11976
• Vector type: Mammalian Expression, Retroviral, CRISPR

Growth in Bacteria

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

Gene/Insert

• Gene/Insert name: Cas9
• Promoter: Ubiquitin
• Tag / Fusion Protein:
  • mCherry (via t2a) (C terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: BamH1 (not destroyed)
• 3′ cloning site: EcoR1 (not destroyed)
• 5′ sequencing primer: Ubi Down
• 3′ sequencing primer: Wre Up

Resource Information

• Supplemental Documents:
  • pRubi-C-T2A-Cas9

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • Takara Bio Limited Use Label License (formerly Clontech)
• Industry Terms:
  • Not Available to Industry

Depositor Comments

mCherry contains an E158D mutation. The depositor states that this mutation should not affect plasmid function.