SpySwitch
(Plasmid #184225)

• Purpose: Expresses SpySwitch in the bacterial cytoplasm. SpySwitch binds SpyTag-, SpyTag002- or SpyTag003-fusions non-covalently for affinity purification, allowing gentle pH or temperature release.
• Depositing Lab: Mark Howarth
• Publication: Vester et al Nat Commun. 2022 Jun 28;13(1):3714. doi: 10.1038/s41467-022-31193-8.

Backbone

• Vector backbone: pDEST14
• Backbone manufacturer: Thermo Fisher Scientific
• Backbone size w/o insert (bp): 3641
• Total vector size (bp): 4055
• Vector type: Bacterial Expression

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: SpySwitch
• Species: Synthetic
• Insert Size (bp): 414
• Mutation: E77A mutation prevents isopeptide bond formation, S49C mutation enables attachment to solid support. K28H, S30H, R32H, S59T, Y84H, E85H, V94I and A111P allow for capture and gentle pH or temperature release of SpyTag fusion proteins.
• Promoter: T7
• Tag / Fusion Protein:
  • His6 (N terminal on insert)

Cloning Information

• Cloning method: Gibson Cloning
• 5′ sequencing primer: T7F
• 3′ sequencing primer: T7R

Terms and Licenses

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