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.
pTagRFP-H2B is a mammalian expression vector encoding TagRFP-H2B fusion protein (see reporter description). The vector can be used for fluorescent labeling of histone H2B in living cells.
Transiently transfected HeLa cells expressing TagRFP fusion with histone H2B.
Image was kindly provided by Michael W. Davidson (Florida State University).
TagRFP codon usage is optimized for high expression in mammalian cells (humanized) [Haas et al., 1996]. Human histone H2B is fused to the TagRFP N-terminus.
pTagRFP-H2B vector can be used as a source of TagRFP-H2B 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
pTagRFP-H2B vector can be transfected into mammalian cells by any known transfection method. CMV promoter provides strong, constitutive expression of the TagRFP-H2B 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
Histone H2B: 657-1034
SV40 early mRNA polyadenylation signal
Polyadenylation signals: 1919-1924 & 1948-1953
mRNA 3' ends: 1957 & 1969
f1 single-strand DNA origin: 2016-2471
Bacterial promoter for expression of Kanr gene
-35 region: 2533-2538
-10 region: 2556-2561
Transcription start point: 2568
SV40 origin of replication: 2812-2947
SV40 early promoter
Enhancer (72-bp tandem repeats): 2645-2716 & 2717-2788
21-bp repeats: 2792-2812, 2813-2833 & 2835-2855
Early promoter element: 2868-2874
Major transcription start points: 2864, 2902, 2908 & 2913
Kanamycin/neomycin resistance gene
Neomycin phosphotransferase coding sequences:
Start codon (ATG): 2996-2998
Stop codon: 3788-3790
G->A mutation to remove Pst I site: 3178
C->A (Arg to Ser) mutation to remove BssH II site: 3524
Herpes simplex virus (HSV) thymidine kinase (TK) polyadenylation signal
Polyadenylation signals: 4026-4031 & 4039-4044
pUC plasmid replication origin: 4375-5018
TagRFP-related materials (also referred to as "Products") are intended for research use only. The Products are covered by U.S. Pat. 7,638,615; 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.
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