pN1-Lck-GCaMP3
(Plasmid #26974)

• Depositing Lab: Baljit Khakh
• Publication: Shigetomi et al Neuron Glia Biol. 2010 Dec 16:1-9. [Epub ahead of print]

Backbone

• Vector backbone: pEGFP-N1
• Backbone manufacturer: Clontech
• Backbone size w/o insert (bp): 3950
• 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: Lck-GCaMP3
• Alt name: GCaMP
• Alt name: GCaMP3
• Alt name: Lck-GCaMP3
• Insert Size (bp): 1480
• Mutation: Plasmid contains N-terminal 26 amino acid membrane targeting sequence of Lck (Src tyrosine kinase) fused to GCaMP3. 6xHis and Xpress (FLAG-like) epitope tags contained within 'RSET' sequence, making up the linker region between Lck and GCaMP3
• Tags / Fusion Proteins:
  • His (N terminal on backbone)
  • Xpress (N terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: BglII (not destroyed)
• 3′ cloning site: NotI (not destroyed)
• 5′ sequencing primer: ACGGTGGGAGGTCTATATAAGCAG AG

Resource Information

• Articles Citing this Plasmid:
  • 6 References

Terms and Licenses

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

Depositor Comments

Sequence is identical to pN1-Lck-GCaMP2 http://www.addgene.org/24794 , except for three mutations in GCaMP2 to make GCaMP3: M153K (EGFP); T203V (EGFP) and N60D (CaM).

Note that the RSET tag refers to the first 37 amino acids of the insert, which include the His tag, T7 tag and Xpress tag: MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVD

A genetically targeted optical sensor to monitor calcium signals in astrocyte processes. Shigetomi E., et al. Nat. Neurosci. 2010 Jun;13(6):759-66.