pACYC-T7-SpCas9-T7-EGFP_gRNA1-T7term (BPK848)
(Plasmid #181745)

• Purpose: dual T7 promoter vector for bacterial expression of SpCas9 with a c-terminal NLS and 3xFLAG tag, along with an SpCas9 sgRNA targeting EGFP site 1
• Depositing Lab: Benjamin Kleinstiver
• Publication: Christie et al Nat Biotechnol. 2022 Oct 6. pii: 10.1038/s41587-022-01492-y. doi: 10.1038/s41587-022-01492-y.

Backbone

• Vector backbone: BPK764 (Addgene Plasmid #65767)
• Vector type: Bacterial Expression ; in vitro transcription; T7 promoter

Growth in Bacteria

• Bacterial Resistance(s): Chloramphenicol, 25 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: Low Copy

Gene/Insert

• Gene/Insert name: human/zebrafish codon optimized SpCas9
• Alt name: BPK848
• gRNA/shRNA sequence: GGGCACGGGCAGCTTGCCGG
• Species: Synthetic
• Promoter: T7
• Tag / Fusion Protein:
  • NLS(SV40)-3xFLAG (C terminal on insert)

Cloning Information

• Cloning method: Gibson Cloning
• 5′ sequencing primer: oBK311-GCCATTCGATGGTGTCCGG
• 3′ sequencing primer: oBK390-GCAAATTCGACCCGGTCGTCG Chloramphenicol

Resource Information

• Supplemental Documents:
  • pACYC-T7-SpCas9-T7-EGFPgRNA1-T7term (BPK848).gb

Terms and Licenses

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