pET24b-APEX2
(Plasmid #111702)

• Purpose: For IPTG-inducible expression of APEX2 in E. coli
• Depositing Lab: Jessica Seeliger
• Publication: Ganapathy et al ACS Infect Dis. 2018 Apr 30. doi: 10.1021/acsinfecdis.8b00044.

Backbone

• Vector backbone: pET24b
• Backbone manufacturer: Novagen
• Total vector size (bp): 6026
• Modifications to backbone: None
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: Low Copy

Gene/Insert

• Gene/Insert name: APEX2
• Insert Size (bp): 819
• Promoter: T7 promoter
• Tags / Fusion Proteins:
  • V5 (N terminal on insert)
  • 6xHis (C terminal on backbone)

Cloning Information

• Cloning method: Ligation Independent Cloning
• 5′ sequencing primer: TAATACGACTCACTATA

Resource Information

• Supplemental Documents:
  • pET24b-APEX2.gb
• Addgene Notes:
  • Addgene Diagnostic Digest
• A portion of this plasmid was derived from a plasmid made by: Alice Ting (pcDNA3-mito-APEX, Addgene plasmid #42607)
• Articles Citing this Plasmid:
  • 2 References

Terms and Licenses

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

Depositor Comments

This plasmid is for IPTG-inducible expression of APEX2 in the E. coli cytoplasm. The APEX gene was subcloned from pcDNA3-mito-APEX and subsequently mutated by site-directed mutagenesis to generate the optimized variant APEX2.

See also references: Rhee HW, Zou P, Udeshi ND, Martell JD, Mootha VK, Carr SA, Ting AY. Proteomic mapping of mitochondria in living cells via spatially restricted enzymatic tagging. Science. 2013;339(6125):1328-31. PMCID: 3916822.

Lam SS, Martell JD, Kamer KJ, Deerinck TJ, Ellisman MH, Mootha VK, Ting AY. Directed evolution of APEX2 for electron microscopy and proximity labeling. Nat Methods. 2015;12(1):51-4. PMCID: 4296904