PurposeExpresses a tandem ddFP heterodimer in mammalian cells, plasmid 1 for a translocation-based red fluorescent caspase-9 biosensor, used together with plasmid 2 (BNLS).
Full plasmid sequence is not available for this item.
|Item||Catalog #||Description||Quantity||Price (USD)|
|Plasmid||60973||Standard format: Plasmid sent in bacteria as agar stab||1||$65|
This material is available to academics and nonprofits only.
- Backbone size w/o insert (bp) 5500
- Total vector size (bp) 7024
Modifications to backboneMulti cloning sites has been replaced by a customized sequence, which is compatible to the reading frame of vector pBAD hisB.
Vector typeMammalian Expression
Selectable markersNeomycin (select with G418)
Growth in Bacteria
Copy numberHigh Copy
Gene/Insert nameddRFP A and ddRFP B
Alt namedimerization dependent red fluorescent protein (ddRFP) copy A
Alt namedimerization dependent red fluorescent protein (ddRFP) copy B (dark copy)
SpeciesSynthetic; synthetic construct
Insert Size (bp)1521
- Promoter CMV
/ Fusion Proteins
- triplicated NLS sequence DPKKKRKV placed after ddRFP A
- NES sequence LALKLAGLDIGS (C terminal on insert)
- Caspase9 substrate sequence (LEHD) is after NLS sequence but before ddFP copy B
- Cloning method Restriction Enzyme
- 5′ cloning site XhoI (not destroyed)
- 3′ cloning site HindIII (not destroyed)
- 5′ sequencing primer T7 Promoter forward primer
- 3′ sequencing primer BGH Reverse primers (Common Sequencing Primers)
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:RANLS-LEHD-BNES was a gift from Robert Campbell (Addgene plasmid # 60973 ; http://n2t.net/addgene:60973 ; RRID:Addgene_60973)
For your References section:Ratiometric biosensors based on dimerization-dependent fluorescent protein exchange. Ding Y, Li J, Enterina JR, Shen Y, Zhang I, Tewson PH, Mo GC, Zhang J, Quinn AM, Hughes TE, Maysinger D, Alford SC, Zhang Y, Campbell RE. Nat Methods. 2015 Jan 26. doi: 10.1038/nmeth.3261. 10.1038/nmeth.3261 PubMed 25622108