refGFP1_pQE30
(Plasmid #48644)

• Purpose: Bact exp vector encoding for the 6XHis tagged insensitive variant of the fluorescent redox probe roGFP with mut Cys147 to Serine, del insertion147b and mutated Cys204 to Glut.Referred to as refGFP
• Depositing Lab: David Ron
• Publication: Avezov et al J Cell Biol. 2013 Apr 15;201(2):337-49. doi: 10.1083/jcb.201211155.

Backbone

• Vector backbone: pQE30
• Backbone size w/o insert (bp): 3455
• Total vector size (bp): 4217
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): M15
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: refGFP1
• Insert Size (bp): 762
• Mutation: refGFP C147S, C204Q
• Tag / Fusion Protein:
  • 6xHis (N terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: BamH1 (unknown if destroyed)
• 3′ cloning site: XhoI (unknown if destroyed)
• 5′ sequencing primer: pQEforward
• 3′ sequencing primer: pQEreverse

Resource Information

• A portion of this plasmid was derived from a plasmid made by: James Remington U. Oregon USA

Terms and Licenses

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

Depositor Comments

This plasmids needs to be amplified and maintained in M15 at all times as this strain has the pREP4 repressor vector that prevents undesired expression (it's important to maintain the plasmid when the protein is not expressed even if it's believed to be inert for the bacteria, to avoid selective pressure in favour of recombinant versions of the gene).