pMLM3705
(Plasmid #47754)

• Purpose: Expresses mammalian cell codon-optimized dCas9-VP64
• Depositing Lab: Keith Joung
• Publication: Maeder et al Nat Methods. 2013 Jul 25. doi: 10.1038/nmeth.2598.

Backbone

• Vector backbone: pJDS246
• Backbone manufacturer: Joung lab (Addgene plasmid # 43861)
• Modifications to backbone: codon-optimized Cas9 modified by site-directed mutagenesis to dCas9; added VP64 activation domain
• Vector type: Mammalian Expression, CRISPR

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): XL1 Blue
• Copy number: Unknown

Gene/Insert

• Gene/Insert name: codon optimized dCas9-VP64
• Alt name: inactive Cas9 nuclease
• Alt name: VP64 activation domain
• Species: Synthetic; Streptococcus pyogenes
• Mutation: D10A and H840A mutations in Cas9 (catalytically inactive)
• Promoter: CMV
• Tags / Fusion Proteins:
  • NLS (C terminal on insert)
  • 3x FLAG (C terminal on insert)
  • VP64 (C terminal on insert)

Cloning Information

• Cloning method: Unknown
• 5′ sequencing primer: CMV-F
• 3′ sequencing primer: BGH-rev

Resource Information

• Articles Citing this Plasmid:
  • 16 References

Terms and Licenses

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

Depositor Comments

Plasmid Features:
Cas9 = 409-4512
NLS = 4519-4539
3X FLAG = 4546-4611
VP64 = 4633-4794