Sa-dCas9-NLS-3xFLAG/pcDNA3.1
(Plasmid #98041)

• Purpose: Expresses Sa-dCas9-NLS-3xFLAG in mammalian cells for enChIP analysis to purify specific genomic regions of interest
• Depositing Lab: Hodaka Fujii
• Publication: Fujita et al BMC Res Notes. 2018 Feb 27;11(1):154. doi: 10.1186/s13104-018-3262-4.

Backbone

• Vector backbone: pcDNA3.1
• Backbone manufacturer: Invitrogen
• Backbone size w/o insert (bp): 5500
• Total vector size (bp): 8767
• Vector type: Mammalian Expression, CRISPR
• Selectable markers: Neomycin (select with G418)

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 30°C
• Growth Strain(s): NEB Stable
• Growth instructions: High frequency of recombination. Culture at 30°C.
• Copy number: Unknown

Gene/Insert

• Gene/Insert name: Sa-dCas9-NLS-3xFLAG
• Species: Synthetic; Staphylococcus aureus
• Insert Size (bp): 3264
• Mutation: D10A + H557A
• Promoter: CMV
• Tags / Fusion Proteins:
  • NLS (nuclear localization signal) (C terminal on insert)
  • 3xFLAG tag (C terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: Nhe I (not destroyed)
• 3′ cloning site: Not I (not destroyed)
• 5′ sequencing primer: T7
• 3′ sequencing primer: BGH Reverse

Resource Information

• A portion of this plasmid was derived from a plasmid made by: A portion of this plasmid was derived from a plasmid made by Keith Joung (Addgene plasmid 70703)
• Article Citing this Plasmid:
  • 1 Reference

Terms and Licenses

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

Depositor Comments

This plasmid can be used to isolate specific genomic regions of interest using a catalytically inactive Cas9 fused with a tag(s).

Purify: locus-specific chromatin immunoprecipitation (enChIP)
This system is compatible with BPK2660 (www.addgene.org/70709) from the Joung lab.

Additional information and protocols can be found at:
http://www.med.hirosaki-u.ac.jp/~bgb/iChIP_protocols/index_e.html

For more information on Fujii Lab CRISPR Plasmids please refer to: http://www.addgene.org/crispr/fujii/