ISceI-GR-RFP
(Plasmid #17654)

• Depositing Lab: Tom Misteli
• Publication: Soutoglou et al Nat Cell Biol. 2007 Jun . 9(6):675-82.

Backbone

• Vector backbone: pDsRed-Monomer-C1
• Backbone manufacturer: Clontech
• Backbone size w/o insert (bp): 4700
• Vector type: Mammalian Expression

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: I-SceI-GRLBD
• Alt name: SCEI
• Alt name: GR ligand binding domain
• Alt name: glucocorticoid receptor
• Species: R. norvegicus (rat), S. cerevisiae (budding yeast)
• Mutation: C655G mutation in the glucocorticoid receptor (see comments)
• Entrez Gene: SCEI (a.k.a. Q0160)
• Entrez Gene: Nr3c1 (a.k.a. GR, Gcr, Grl)
• Tag / Fusion Protein:
  • mRFP (N terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: SacI (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: DsRed1-C
• 3′ sequencing primer: SV40pA-R

Resource Information

• Articles Citing this Plasmid:
  • 12 References

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • Takara Bio Limited Use Label License (formerly Clontech)
• Industry Terms:
  • Not Available to Industry

Depositor Comments

The Ligand Binding
Domain (LBD) of GR was amplified from pCI-nGFP-C656G GR (gift from G. Hager) and cloned into pEGFP-C2
(Clontech) in the SacII and BamHI sites. The C656G mutation of GR that makes it more responsive to ligand and
trasnfers it more quickly to the nucleus. Similarly, the I-SceI cDNA was amplified from HA-I-SceI and cloned into the previously described
GFP-GRLBD into the SacI and SalI restriction sites. Finally, the chimeric I-SceI–GR cDNA was subcloned into pDsRed-Monomer-C1 (Clontech) into the SacI and BamHI sites.