pTRE-T2-∆mIR-IRES-Puromycin
(Plasmid #105168)

• Purpose: (Empty Backbone) Mammalian expression vector for expressing tandem cDNAs
• Depositing Lab: Michael E. Wright
• Publication: Michael E Wright Lab Plasmids (unpublished)

Backbone

• Vector backbone: pTRE-T2-mIR-Puromycin
• Backbone manufacturer: Clontech = pTRE-T2
• Backbone size (bp): 4847
• Vector type: Mammalian Expression
• Promoter: Tet/CMV
• Selectable markers: Puromycin

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Growth instructions: Methylation deficient cells from NEB to alleviate methylation sensitive restriction sites like Xba I. However not recommended for long-term propagation and storage.
• Copy number: High Copy

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ sequencing primer: AGG CGT ATC ACG AGG CCC TTT CGT
• 3′ sequencing primer: TAT TAC CGC CTT TGA GTG AGC TGA

Terms and Licenses

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

Depositor Comments

A synthetic DNA containing the IRES element was cloned into EcoRI/Xba I site of pTRE-T2-miR-Puromycin vector after removal of the mIR cassette. First digest parental pTRE-T2-miR-Puromycin vector with EcoRI and Xba I to remove the miR cassette. Then the IRES cassette was ligated back to the EcoRI site and Xba I site. The IRES cassette contains the unique EcoRI site followed by unique NotI and Mlu I sites that is followed by the start of the IRES sequence. This cloning site will allow insertion of the 1st cDNA. On the 3’ end of IRES there are unique Spe I and Xba I cloning sites for insertion of the second cDNA into the cassette.

Please note- to use the BsaI and XbaI sites for cloning, you must use unmethylated DNA. You will need to isolate the DNA from a methylation deficient bacterial strain.