hTRIM65_EGFP_N1
(Plasmid #54320)

• Purpose: Mammalian expression vector for human TRIM65 fusion protein with c-terminal EGFP.
• Depositing Lab: Martin Dorf
• Publication: Li et al Proc Natl Acad Sci U S A. 2014 May 13;111(19):6970-5. doi: 10.1073/pnas.1322545111. Epub 2014 Apr 28.

Backbone

• Vector backbone: pEGFP-N1
• Backbone manufacturer: Clontech
• Backbone size w/o insert (bp): 4723
• Total vector size (bp): 6281
• Modifications to backbone: human TRIM65 in inserted into pEGFP-N1 with 5': XhoI/SalI insertion + kozak initiation sequence, and 3': EcoRI insertion. The stop codon is removed, and one addidional C nucleotide is added before the EcoRI site to maintain the reading frame. The construct contains : (partial XhoI site) CTCGACGCCACC-human TRIM65 CDS(1551bp, no stop)-C-(GAATTC EcoRI site)-44bp-EGFP.
• 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: High Copy

Gene/Insert

• Gene/Insert name: TRIM65
• Alt name: tripartite motif containing 65
• Species: H. sapiens (human)
• Insert Size (bp): 1558
• GenBank ID: BC021259.2 (has same CDS sequence)
• Entrez Gene: TRIM65 (a.k.a. 4732463G12Rik)
• Promoter: CMV
• Tag / Fusion Protein:
  • EGFP (C terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: XhoI (destroyed during cloning)
• 3′ cloning site: EcoRI (not destroyed)
• 5′ sequencing primer: CMV forward (cgtgtacggtgggaggtct)
• 3′ sequencing primer: EGFP reverse (tccagctcgaccaggatg)

Terms and Licenses

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

Depositor Comments

Human TRIM65 cDNA was cloned from the T98 glioblastoma cell line (ATCC). The CDS (coding sequence) is identical to Genbank BC021259.2.