GPD-roGFP
(Plasmid #49436)

• Purpose: Expresses thiol redox-sensitive ratiometric sensor roGFP in the mitochondrial intermembrane space of mammalian cells
• Depositing Lab: Paul Schumacker
• Publication: Waypa et al Circ Res. 2010 Feb 19;106(3):526-35. doi: 10.1161/CIRCRESAHA.109.206334. Epub 2009 Dec 17.

Backbone

• Vector backbone: VQ Ad5CMV K-NpA
• Backbone manufacturer: ViraQuest Inc.
• Backbone size w/o insert (bp): 5217
• Total vector size (bp): 6200
• Modifications to backbone: None
• Vector type: Mammalian Expression, Adenoviral

Growth in Bacteria

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

Gene/Insert

• Gene/Insert name: GPD-roGFP2
• Alt name: IMS-roGFP
• Species: Synthetic
• Insert Size (bp): 3150
• Mutation: C48S, Q80R, S147C, and Q204C in Clontech's GFP
• Promoter: CMV
• Tag / Fusion Protein:
  • Glycerol phosphate dehydrogenase, 5' to roGFP2 (N terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: XhoI (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: 5'GGCGTGTACGGTGGGAGGTCTATA-3'
• 3′ sequencing primer: 5'-TTAAAAAACCTCCCCACCTCCCCC-3'

Resource Information

• Supplemental Documents:
  • Description of GPD-roGFP2
• A portion of this plasmid was derived from a plasmid made by: We inserted mutations into Clontech's eGFP gene to convert it into roGFP, and cut the roGFP from the Clontech backbone. We ligated cDNA encoding glycerol phosphate dehydrogenase (GPD)(5') to the roGFP dDNA (3') - a ClaI site separates the GPD and roGFP2 genes. The GPD-roGFP construct was ligated into the VQ vector between XhoI (5') and BamHI (3') sites.
• Articles Citing this Plasmid:
  • 4 References

Terms and Licenses

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