PurposeExpression of a GFP-dependent transcription factor component in mammalian cells
|Item||Catalog #||Description||Quantity||Price (USD)|
|Plasmid||49438||Standard format: Plasmid sent in bacteria as agar stab||1||$75|
This material is available to academics and nonprofits only.
- Backbone size w/o insert (bp) 4823
- Total vector size (bp) 5666
Vector typeMammalian Expression
Growth in Bacteria
Copy numberHigh Copy
Gene/Insert nameGBP1-Gal4 DNA binding domain fusion protein
Insert Size (bp)843
MutationC98S (to increase protein solubility), Q3D
- Promoter CAG
- Cloning method Restriction Enzyme
- 5′ cloning site AgeI (not destroyed)
- 3′ cloning site NotI (not destroyed)
- 5′ sequencing primer GGACTTCCTTTGTCCCAAATCTG
- 3′ sequencing primer TAGCCAGAAGTCAGATGCTC (Common Sequencing Primers)
The GBP1 used differs from the PDB GBP1 by two amino acids. One is Q3D at a site away from the GFP binding pocket and not expected to affect GFP binding. The second was a C98S mutation to enhance protein solubility.
Kirchhofer, A. et al. (2010). Modulation of protein properties in living cells using nanobodies. Nat. Struct. Mol. Biol. 17, 133–138.
Tang J.C. et al. (2013). A nanobody-based system using fluorescent proteins as scaffolds for cell-specific gene manipulation. Cell. 2013 Aug 15;154(4):928-39.
These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the plasmids were described, and include Addgene in the Materials and Methods of your future publications.
For your Materials & Methods section:pCAG-GBP1-10gly-Gal4DBD was a gift from Connie Cepko (Addgene plasmid # 49438 ; http://n2t.net/addgene:49438 ; RRID:Addgene_49438)
For your References section:A nanobody-based system using fluorescent proteins as scaffolds for cell-specific gene manipulation. Tang JC, Szikra T, Kozorovitskiy Y, Teixiera M, Sabatini BL, Roska B, Cepko CL. Cell. 2013 Aug 15;154(4):928-39. doi: 10.1016/j.cell.2013.07.021. 10.1016/j.cell.2013.07.021 PubMed 23953120