pCX-TRI-2A
(Plasmid #74674)

• Purpose: mammalian expression of TRI-2A (hHO1/F2A/hCD73/F2A/hCD39)
• Depositing Lab: Roberto Giovannoni
• Publication: Cinti et al PLoS One. 2015 Oct 29;10(10):e0141933. doi: 10.1371/journal.pone.0141933. eCollection 2015.

Backbone

• Vector backbone: pCX-2A-2A
• Backbone manufacturer: Giovannoni lab, Addgene plasmid 74670
• Total vector size (bp): 10110
• 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: Unknown

Gene/Insert 1

• Gene/Insert name: HO1
• Species: H. sapiens (human)
• Entrez Gene: HMOX1 (a.k.a. HMOX1D, HO-1, HSP32, bK286B10)
• Promoter: pCAGGS
• Tag / Fusion Protein:
  • F2A (C terminal on backbone)

Cloning Information for Gene/Insert 1

• Cloning method: Restriction Enzyme
• 5′ cloning site: EcoRI (not destroyed)
• 3′ cloning site: NheI (not destroyed)
• 5′ sequencing primer: pCAX-F CAGCTCCTGGGCAACGTGC

Gene/Insert 2

• Gene/Insert name: CD73
• Species: H. sapiens (human)
• Entrez Gene: NT5E (a.k.a. CALJA, CD73, E5NT, NT, NT5, NTE, eN, eNT)
• Tag / Fusion Protein:
  • F2A (C terminal on backbone)

Cloning Information for Gene/Insert 2

• Cloning method: Restriction Enzyme
• 5′ cloning site: AflII (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: none

Gene/Insert 3

• Gene/Insert name: CD39
• Species: H. sapiens (human)
• Entrez Gene: ENTPD1 (a.k.a. ATP-DPH, ATPDase, CD39, NTPDase-1, SPG64)

Cloning Information for Gene/Insert 3

• Cloning method: Restriction Enzyme
• 5′ cloning site: XhoI (not destroyed)
• 3′ cloning site: EcoRI (not destroyed)
• 5′ sequencing primer: none
• 3′ sequencing primer: M13R

Resource Information

• A portion of this plasmid was derived from a plasmid made by: HMOX1 (hHO-1) sequence was kindly provided by Prof. Fritz Bach's laboratory at Harvard Medical School, Boston. NT5E (hCD73) sequence was kindly provided by Dr. Jozef Spychala at University of Michigan, Ann Arbor. ENTPD1 (hCD39) sequence was kindly provided by Dr. Simon Robson at Harvard Medical School, Boston.

Terms and Licenses

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

Depositor Comments

Contains a D473N mutation in CD73, but it does not impair either the plasmid function or the hCD73 protein activity.