lentiCas9-EGFP
(Plasmid #63592)

• Purpose: Expresses human codon-optimized S. pyogenes Cas9 protein and EGFP from EFS promoter. 3rd generation lentiviral backbone.
• Depositing Labs: Phil Sharp, Feng Zhang
• Publication: Chen et al Cell. 2015 Mar 12;160(6):1246-60. doi: 10.1016/j.cell.2015.02.038. Epub 2015 Mar 5.

Backbone

• Vector backbone: pFUGW
• Total vector size (bp): 13178
• Vector type: Mammalian Expression, Lentiviral, CRISPR

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): Stbl3
• Growth instructions: Use SapI digest to check for unwanted recombination of lentiviral plasmid. Only amplify in RecA- bacteria (eg. Stbl3).
• Copy number: High Copy

Gene/Insert 1

• Gene/Insert name: Cas9
• Alt name: S. pyogenes CRISPR-Cas9
• Species: Synthetic
• Insert Size (bp): 4200
• Promoter: EFS-NS
• Tag / Fusion Protein:
  • FLAG (C terminal on insert)

Cloning Information for Gene/Insert 1

• Cloning method: Restriction Enzyme
• 5′ cloning site: AgeI, XbaI, AfeI (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: pLLU-F
• 3′ sequencing primer: EGFP-N

Gene/Insert 2

• Gene/Insert name: EGFP
• Promoter: EFS-NS
• Tag / Fusion Protein:
  • P2A (N terminal on insert)

Cloning Information for Gene/Insert 2

• Cloning method: Unknown
• 5′ sequencing primer: needs custom
• 3′ sequencing primer: WPRE-R

Resource Information

• Supplemental Documents:
  • lentiCas9EGFP genbank file
• Articles Citing this Plasmid:
  • 40 References

Terms and Licenses

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

Depositor Comments

Special note from the Zhang lab: We are constantly improving our CRISPR reagents. Please check https://zlab.bio/ for the most up-to-date information.