pRM053 His-TEV-Cas9 triNLS 0C
(Plasmid #196244)

• Purpose: Expression of NLS-rich Cas9 (0 Cys residues, mammalian codon optimized)
• Depositing Lab: Ross Wilson
• Publication: Foss et al Nat Biomed Eng. 2023 May;7(5):647-660. doi: 10.1038/s41551-023-01032-2. Epub 2023 Apr 25.

Backbone

• Vector backbone: pEC-K-MBP
• Backbone manufacturer: pET-based custom vector
• Backbone size w/o insert (bp): 6400
• Total vector size (bp): 9073
• Modifications to backbone: MBP removed
• Vector type: Bacterial Expression, CRISPR

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: Cas9
• Alt name: Cas9-0C-triNLS
• Species: Streptococcus pyogenes
• Insert Size (bp): 4102
• Mutation: Cys at positions 80 and 574 have been mutated to Ser
• Tags / Fusion Proteins:
  • His6 (N terminal on backbone) (N terminal on insert)
  • HA (C terminal on insert)
  • SV40 (C terminal on insert)
  • Nucleoplasmin (C terminal on insert)
  • NLS (N terminal on insert)

Cloning Information

• Cloning method: Ligation Independent Cloning
• 5′ sequencing primer: T7 promoter
• 3′ sequencing primer: T7 terminator

Resource Information

• Supplemental Documents:
  • pRM053_02.gbk
• A portion of this plasmid was derived from a plasmid made by: pMJ806 was a gift from Jennifer Doudna (Addgene plasmid # 39312 ; http://n2t.net/addgene:39312 ; RRID:Addgene_39312)

Terms and Licenses

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