G-GECO1.2-Orai1
(Plasmid #73562)

• Purpose: Green fluorescent reporter of Orai1-associated calcium influx
• Depositing Lab: Michael Cahalan
• Publication: Dynes et al Proc Natl Acad Sci U S A. 2016 Jan 12;113(2):440-5. doi: 10.1073/pnas.1523410113. Epub 2015 Dec 28.

Backbone

• Vector backbone: pEGFP-C2
• Backbone manufacturer: Clontech
• Backbone size w/o insert (bp): 3954
• Total vector size (bp): 6138
• Modifications to backbone: The EGFP gene has been replaced with the genetically encoded calcium indicator G-GECO1.2
• Vector type: Mammalian Expression
• Selectable markers: Neomycin (select with G418)

Growth in Bacteria

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

Gene/Insert

• Gene/Insert name: G-GECO1.2-Orai1
• Species: H. sapiens (human), Synthetic
• Insert Size (bp): 2184
• GenBank ID: NM_032790.3 JN258415.1
• Entrez Gene: ORAI1 (a.k.a. CRACM1, IMD9, ORAT1, TAM2, TMEM142A)
• Promoter: CMV Immediate Early
• Tag / Fusion Protein:
  • G-GECO1.2 (N terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: AgeI (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: TGACGTCAATGGGAGTTTG
• 3′ sequencing primer: ACATTGATGAGTTTGGAC

Resource Information

• Articles Citing this Plasmid:
  • 4 References

Terms and Licenses

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

Depositor Comments

The fusion protein in this plasmid contains the genetically encoded calcium indicator G-GECO1.2 from the plasmid CMV-G-GECO1.2 (Addgene plasmid # 32446) provided by Robert Campbell. Note that the hOrai1 gene isolated in the Cahalan lab contains a silent mutation/polymorphism (T798C; aa T266).