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URL: http://www.addgene.org/14860
Proper Citation: RRID:Addgene_14860
Insert Name: C kinase activity reporter
Bacterial Resistance: Ampicillin
Defining Citation: PMID:12782683
Vector Backbone Description: Backbone Marker:Invitrogen; Backbone Size:5400; Vector Backbone:pcDNA3; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Comments: PKC substrate sequence flanked by flexible linker sequence: GGSGG RFRRFQTLKIKAKA GGSGG. A206K mutation present in both CFP (mCFP) and citrine (mYFP) to reduce the intrinsic homoaffinity of all GFPs and preclude intermolecular FRET by CFP-YFP dimerization. CFP was amplified by PCR from a plasmid template to encode a HindIII restriction site followed by a consensus initiation site for translation (CGCCACC) before the initiating ATG of CFP, and a KpnI restriction site at the 3' end instead of a terminating codon. FHA2, a gift from Michael Yaffe (MIT, Cambridge, MA) was amplified by PCR to include KpnI and BamHI restriction sites at the 5' and 3' ends, respectively. Citrine was amplified by PCR to include a 5' BamHI followed by the PKC substrate sequence and a 3' XbaI following the terminating codon.
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Source: Addgene
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