murine TCR OTI-2A.pMIG II
(Plasmid #52111)

• Purpose: retroviral expression of both TCRalpha and TCRbeta using a single vector
• Depositing Lab: Dario Vignali
• Publication: Holst et al Nat Protoc. 2006;1(1):406-17.

Backbone

• Vector backbone: pMIG II
• Backbone manufacturer: Dario Vignali, Addgene plasmid 52107
• Total vector size (bp): 8155
• Vector type: Mammalian Expression, Retroviral

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): Stbl3
• Copy number: High Copy

Gene/Insert 1

• Gene/Insert name: T cell receptor alpha chain
• Alt name: Valpha2
• Species: M. musculus (mouse)
• Entrez Gene: Tcra (a.k.a. Tcralpha)
• Tag / Fusion Protein:
  • T2A (from Thosea asigna virus) (C terminal on insert)

Cloning Information for Gene/Insert 1

• Cloning method: Unknown
• 5′ sequencing primer: pLXSN5

Gene/Insert 2

• Gene/Insert name: T cell receptor beta chain
• Alt name: Vbeta5.2
• Species: M. musculus (mouse)
• Entrez Gene: Tcrb (a.k.a. TCRbeta, Tib)

Cloning Information for Gene/Insert 2

• Cloning method: Unknown
• 5′ sequencing primer: na
• 3′ sequencing primer: EGFP-N

Resource Information

• Articles Citing this Plasmid:
  • 4 References

Terms and Licenses

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

Depositor Comments

The cDNAs encoding the TCRa (lacking the stop codon) and TCRb chains were linked via a 2A peptide into a single open reading frame by recombinant PCR and cloned into a mouse stem cell virus (MSCV)-based retroviral vector containing an IRES-GFP cassette to facilitate the identification of marked cells.
The 2A peptide remains attached to the TCRa carboxy terminus and can be used to verify the complete separation of the two chains. Inclusion of a Gly-Ser-Gly (GSG) spacer between TCRa and the 2A peptide ensures complete ‘cleavage’. We have not seen any deleterious effects of the 2A peptide on the function of any proteins examined thus far.