pCAG-CalfluxCTN
(Plasmid #190068)

• Purpose: Expresses BRET Ca2+ sensor CalfluxCTN in mammalian cells
• Depositing Lab: Carl Johnson
• Publication: Cumberbatch et al Sci Signal. 2022 Aug 16;15(747):eabq7618. doi: 10.1126/scisignal.abq7618. Epub 2022 Aug 16.

Backbone

• Vector backbone: pcDNA3.1/Puro-CAG-VSFP-CR
• Backbone manufacturer: Dr. Michael Lin and his colleagues
• Backbone size w/o insert (bp): 6075
• Total vector size (bp): 7532
• Modifications to backbone: pcDNA3.1/Puro-CAG-VSFP-CR (Addgene ID#: 40257) was used as the backbone, where the VSFP portions were removed and CalfluxCTN coding sequence was inserted at the NheI and BamHI restriction enzyme sites.
• Vector type: Mammalian Expression
• Selectable markers: Puromycin

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: CalfluxCTN
• Species: Synthetic
• Insert Size (bp): 1449
• Promoter: CAG promoter

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: NheI (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: pCAG-F (5'-GCAACGTGCTGGTTATTGTG-3')
• 3′ sequencing primer: BGH-rev (5'-TAGAAGGCACAGTCGAGG-3')

Resource Information

• Supplemental Documents:
  • pCAG-CalfluxCTN.gb

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • NanoLuc Luciferase Limited Use Label License
• Industry Terms:
  • Not Available to Industry

Depositor Comments

CalfluxCTN BRET-based calcium sensor composed of the following 5 segments: Segment 1: Clover C9 - Amino acid residues 1-230 of Clover, Segment 2: Linker - MQVAD, Segment 3: TnC - EF-hand motif derived from Opsanus tau troponin C, Segment 4: Linker - IYPEL, Segment 5: NanoLuc - NanoLuc luciferase