The vector sequence has been compiled using the information from sequence databases, published literature, and other sources, together with partial sequences obtained by Evrogen. This vector has not been completely sequenced.
pFusionRed-cadherin is a mammalian expression vector encoding FusionRed-VE-cadherin fusion protein (see reporter description). The vector can be used for fluorescent labeling of cadherin in living cells.
HeLa cell expressing fusion of human VE-cadherin with FusionRed.
Scale bar represents 10 μm. Data from Shemiakina et al., 2012
FusionRed codon usage is optimized for high expression in mammalian cells (humanized) [Haas et al., 1996]. Human VE-cadherin is fused to the FusionRed N-terminus.
pFusionRed-cadherin vector can be used as a source of FusionRed-VE-cadherin hybrid sequence. The vector backbone contains unique restriction sites that permit its excision and further insertion into expression vector of choice.
Note: The plasmid DNA was isolated from dam+-methylated
The vector backbone contains immediate early promoter of cytomegalovirus (PCMV IE) for protein expression, SV40 origin for replication in mammalian cells expressing SV40 T-antigen, pUC origin of replication for propagation in
SV40 early promoter (PSV40) provides neomycin resistance gene (Neor) expression to select stably transfected eukaryotic cells using G418. Bacterial promoter (P) provides kanamycin resistance gene expression (Kanr) in
Expression in mammalian cells
pFusionRed-cadherin vector can be transfected into mammalian cells by any known transfection method. CMV promoter provides strong, constitutive expression of the FusionRed-VE-cadherin fusion in eukaryotic cells. If required, stable transformants can be selected using G418 [Gorman, 1985].
Suitable host strains for propagation in
Location of features
PCMV IE: 1-589
Enhancer region: 59-465
TATA box: 554-560
Transcription start point: 583
Kozak consensus translation initiation site: 597-607
VE-Cadherin-FusionRed fusion: 604-3684
Start codon (ATG): 604-606
Last amino acid in VE-Cadherin: 2953-2955
Stop codon: 3682-3684
SV40 early mRNA polyadenylation signal
Polyadenylation signals: 3837-3842 & 3866-3871
mRNA 3' ends: 3875 & 3887
f1 single-strand DNA origin: 3934-4389
Bacterial promoter for expression of Kanr gene
-35 region: 4451-4456
-10 region: 4474-4479
Transcription start point: 4486
SV40 origin of replication: 4730-4865
SV40 early promoter
Enhancer (72-bp tandem repeats): 4563-4634 & 4635-4706
21-bp repeats: 4710-4730, 4731-4751 & 4753-4773
Early promoter element: 4786-4792
Major transcription start points: 4782, 4820, 4826 & 4831
Kanamycin/neomycin resistance gene
Neomycin phosphotransferase coding sequences:
Start codon (ATG): 4914-4916
Stop codon: 5706-5708
G->A mutation to remove Pst I site: 5096
C->A (Arg to Ser) mutation to remove BssH II site: 5442
Herpes simplex virus (HSV) thymidine kinase (TK) polyadenylation signal
Polyadenylation signals: 5944-5949 & 5957-5962
pUC plasmid replication origin: 6293-6936
FusionRed-related materials (also referred to as "Products") are intended for research use only. The Products are covered by European Pat. 1994149 and other Evrogen Patents and/or Patent applications pending. By use of these Products, you accept the terms and conditions of the applicable Limited Use Label License.
The CMV promoter is covered under U.S. Patents 5,168,062 and 5,385,839, and its use is permitted for research purposes only. Any other use of the CMV promoter requires a license from the University of Iowa Research Foundation, 214 Technology Innovation Center, Iowa City, IA 52242.
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