pEYFPC1-ECFP/GgVcl 1-419 control 1 FRET
(Plasmid #46277)

• Depositing Lab: Susan Craig
• Publication: Chen et al J Cell Biol. 2005 May 9;169(3):459-70.

Backbone

• Vector backbone: pEYFP-C1
• Backbone manufacturer: Clonetech
• Backbone size w/o insert (bp): 4700
• Total vector size (bp): 6732
• Vector type: Mammalian Expression
• Selectable markers: Neomycin (select with G418)

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: Unknown

Gene/Insert

• Gene/Insert name: ECFP + Vcl 1-419
• Alt name: Vinculin
• Alt name: Vcl
• Species: G. gallus (chicken)
• Mutation: aa 1-419
• Entrez Gene: VCL (a.k.a. VINC1)
• Promoter: CMV
• Tag / Fusion Protein:
  • EYFP (N terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: HindIII (not destroyed)
• 3′ cloning site: KpnI (not destroyed)
• 5′ sequencing primer: CMV-F
• 3′ sequencing primer: SV40pA-R

Terms and Licenses

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

Depositor Comments

To make the control FRET probe pEYFP-C1/CFPV1-400, a HindIII site was engineered before the ATG site of pET15b/CFP-V1-851 and a KpnI site after codon 400 by PCR amplification with 5-CAAGCTTCGatggtgagcaagggc-3 and 5-GGTACCTCAtgcaactttccttgc-3. The PCR product was introduced into TOPO pCRII and sequenced. The HindIII–KpnI fragment of CFP-vinculin1-400 was subcloned into pEYFP-C1 to generate the control plasmid pEYFP-CFPV1-400.

Tandem EYFP-ECFP- hooked to portion of vinculin that localizes to FA; maximum FRET.