pet16b-Avi-ubiquitin_rbs_BirA
(Plasmid #134897)

• Purpose: Expresses HIS-Avi-tagged ubiquitin for recombinant protein production
• Depositing Lab: Andrew Deans
• Publication: Tan et al PLoS One. 2020 Feb 24;15(2):e0229000. doi: 10.1371/journal.pone.0229000. eCollection 2020.

Backbone

• Vector backbone: pET16b
• Backbone manufacturer: Novagen
• Backbone size w/o insert (bp): 5711
• Total vector size (bp): 6984
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Growth instructions: Need to retransform to BL21-DE3 or other suitable strain for protein expression
• Copy number: Low Copy

Gene/Insert 1

• Gene/Insert name: Ubiquitin
• Species: H. sapiens (human)
• Insert Size (bp): 339
• GenBank ID: 7314
• Entrez Gene: UBB (a.k.a. HEL-S-50)
• Promoter: T7
• Tag / Fusion Protein:
  • 10HIS-AviTag-PreScission(3C)protease (N terminal on insert)

Cloning Information for Gene/Insert 1

• Cloning method: Restriction Enzyme
• 5′ cloning site: NcoI (not destroyed)
• 3′ cloning site: EcoRV (destroyed during cloning)
• 5′ sequencing primer: TAATACGACTCACTATAG

Gene/Insert 2

• Gene/Insert name: BirA
• Alt name: Biotin Ligase
• Species: E.coli
• Insert Size (bp): 966
• GenBank ID: U00006.1
• Promoter: RBS

Cloning Information for Gene/Insert 2

• Cloning method: Restriction Enzyme
• 5′ cloning site: EcoRV (destroyed during cloning)
• 3′ cloning site: XhoI (not destroyed)
• 5′ sequencing primer: T7 promoter
• 3′ sequencing primer: T7 terminator primer

Resource Information

• Supplemental Documents:
  • pET16b-Avi-ubiquitin_rbs_BirA.gb
  • Protocol for production of biotinylated Ubiquitin in E.coli

Terms and Licenses

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