pcDNA3.1 Flag YFP hNKCC1 HA-ECL4 (NT935)
(Plasmid #49065)

• Purpose: Expresses human NKCC1 mutant with HA tag inserted into extracellular loop #4 and N-terminal 3xFLAG-YFP tag in mammalian cells. cDNA is synthetic, containing convenient restriction sites.
• Depositing Lab: Biff Forbush
• Publication: Somasekharan et al PLoS One. 2013 Dec 5;8(12):e82060. doi: 10.1371/journal.pone.0082060.

Backbone

• Vector backbone: pcDNA3.1 neo
• Backbone manufacturer: Life Technologies
• Backbone size w/o insert (bp): 5316
• Vector type: Mammalian Expression
• Selectable markers: Neomycin (select with G418)

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: human NKCC1 (synthetic)
• Alt name: Slc12A2 (synthetic)
• Alt name: Na-K-Cl cotransporter
• Species: H. sapiens (human), Synthetic
• Insert Size (bp): 4536
• Mutation: 2xHA epitope in ECL4 inserted at aa574 in hNKCC1 in NT17
• Entrez Gene: SLC12A2 (a.k.a. BSC, BSC-2, BSC2, CCC1, KILQS, NKCC1, PPP1R141)
• Promoter: CMV
• Tags / Fusion Proteins:
  • 3x Flag (N terminal on insert)
  • mVenus (N terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: NheI (not destroyed)
• 3′ cloning site: PmeI (not destroyed)
• 5′ sequencing primer: CMV-F
• 3′ sequencing primer: BGH-rev

Terms and Licenses

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

Depositor Comments

Plasmid constructed from pcDNA3.1 Flag YFP hNKCC1 WT (NT17). See Addgene plasmid# 49085 (http://www.addgene.org/49085/) for plasmid map of wild-type construct.