RG6 GLS Intron 14 del
(Plasmid #110394)

• Purpose: RG6 Fluorescent protein splicing reporter with human GLS intron 14 sequence on first intron space, deleted for putative HuR binding site
• Depositing Lab: Sandra Martha Gomes Dias
• Depositing Organization: Centro Nacional de Pesquisa em Energia e Materiais (CNPEM)
  Located in Brazil
• Publication: HuR (ELAVL1) silencing sensitizes triple-negative breast cancer to glutaminase-targeted therapy (unpublished)

Backbone

• Vector backbone: RG6
• Backbone manufacturer: #80167
• Backbone size w/o insert (bp): 8137
• Total vector size (bp): 8582
• Vector type: Mammalian Expression ; Putative HuR binding site mutated
• Selectable markers: Neomycin (select with G418)

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: dsRed and GFP
• Species: Synthetic
• Mutation: Putative HuR binding site deleted
• Promoter: CMV
• Tag / Fusion Protein:
  • FLAG (N terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: XbaI (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: RG6f 5'-CAAAGTGGAGGACCCAGTACC-3'
• 3′ sequencing primer: RG6r 5'-GCGCATGAACTCCTTGATGAC-3'

Terms and Licenses

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

Depositor Comments

First intron from original RG6 (Dr. Thomas Cooper, Plasmid #80167) was removed (using XbaI*) and replaced by GLS intron 14 (from pGEMT easy Intron 14, Addgene #110397).

Mutagenesis with primers 5'-AAGAATTATGCAAAACCAACAG-3' and 5'-TTCTACTTAGAAGTTCTTCATATTATC-3'

*XbaI site is dam-blocked. Use dam defective strain.