PX458_3XHA_den1FnCas9_KRAB_T2A_eGFP
(Plasmid #200910)

• Purpose: 3XHA tagged dead en1Cas9 from F. novicida with KRAB repressor domain, T2A self-cleaving peptide and enhanced GFP biomarker for mammalian expression
• Depositing Lab: Debojyoti Chakraborty
• Publication: 3XHA tagged dead en1Cas9 from F. novicida with KRAB repressor domain, T2A self-cleaving peptide region and enhanced GFP biomarker for mammalian expression (unpublished)

Backbone

• Vector backbone: PX458
• Backbone size w/o insert (bp): 4832
• Total vector size (bp): 10320
• Vector type: Mammalian 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: 3XHA_den1FnCas9_KRAB_eGFP
• Alt name: Nuclease-dead en1FnCas9 with 3XHA tag, KRAB domain and eGFP for mammalian expression
• Alt name: en1FnCas9 is FnCas9b with E1369R substitution
• Species: Synthetic; Francisella novicida U112
• Insert Size (bp): 5488
• Mutation: Changed Glutamic Acid to Arginine at 1369 position
• GenBank ID: NZ_CP009607.1
• Promoter: CBH
• Tags / Fusion Proteins:
  • 3xHA-SV40NLS (N terminal on insert)
  • Nucleoplasmin NLS-KRAB-T2A-EGFP (C terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: BamH1 (not destroyed)
• 3′ cloning site: Eco53kI (not destroyed)
• 5′ sequencing primer: CTGGCTTATAGGTCTAGGCGCGTGAAGATCAAAAG
• 3′ sequencing primer: CTTTTGATCTTCACGCGCCTAGACCTATAAGCCAG

Resource Information

• A portion of this plasmid was derived from a plasmid made by: Subcloned from PX458, a gift from Feng Zhang's Lab

Terms and Licenses

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