pECIA2
(Plasmid #47032)

• Purpose: (Empty Backbone) This is a Gateway destination vector for secreted expression in Drosophila cell culture by induction via CuSO4. Inserts will be C-terminally tagged with Fc, V5 and hexahistidine tags.
• Depositing Lab: Chris Garcia
• Publication: Ozkan et al Cell. 2013 Jul 3;154(1):228-39. doi: 10.1016/j.cell.2013.06.006.

Backbone

• Vector backbone: pMT/BiP/V5/His A
• Backbone manufacturer: Invitrogen
• Backbone size (bp): 4349
• Vector type: Insect Expression ; Secreted expression in Drosophila Culture
• Promoter: Metallothionein (Copper-Inducible)
• Tags / Fusion Proteins:
  • HRV 3C Protease Cleavage site (C terminal on backbone)
  • Fc(human IgG1) (C terminal on backbone)
  • V5 (C terminal on backbone)
  • Hexahistidine (C terminal on backbone)

Growth in Bacteria

• Bacterial Resistance(s): Chloramphenicol and Ampicillin, 25 & 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): ccdB Survival
• Copy number: High Copy

Cloning Information

• Cloning method: Gateway Cloning
• 5′ sequencing primer: MT-Upstream

Resource Information

• Supplemental Documents:
  • genbank file

Terms and Licenses

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

Depositor Comments

Inducible with 0.8 uM Copper Sulfate. Targets proteins to the extracellular millieu using the Drosophila melanogaster BiP signal peptide. Expresses in Drosophila melanogaster cell lines, such as Schneider 2 (S2) cells. Published in Ozkan et al, Cell 154(1): 228 (2013); PMID: 23827685; doi: 10.1016/j.cell.2013.06.006

After recombination, transform and grow in DH5alpha. The plasmid is no longer chloramphenicol resistant after recombination.