pKillerRed-Mem

Catalog No. PVT10851
Packing 2ug

 

pKillerRed-Mem Information

Function Mammal reporter plasmid

Promoter: CMV promoter

Replicator: Ori, F1 ori, SV40 ori

Terminator: SV40 poly (A) signal

Plasmid classification: lactation cell plasmids; lactating reporter plasmid; fluorescent localization plasmid

Plasmid size: 4787bp

Plasmid label: C-mem, C-KillerRed

Prokaryotic resistance: kanamycin Kan

Screening markers: neomycin Neo/G418

Cloned strain: Escherichia coli DH5 alpha

Culture conditions: 37 centigrade, aerobic, LB

Expression host: mammalian cells such as 293T

 

pKillerRed-Mem Description

PKillerRed-mem is a plasmid which is red fluorescence located in cell membrane of mammalian cells.

pKillerRed-mem is a mammalian expression vector encoding membrane-targeted KillerRed. KillerRed localized on cellular membrane can be used for effective light-induced cell killing.KillerRed codon usage is optimized for high expression in mammalian cells (humanized) [Haas et al. 1996]. Membrane localization signal (MLS) of neuromodulin is linked to the KillerRed N-terminus. The MLS (Nterminal 20 amino acid residues of neuromodulin) contains a signal for posttranslational palmitoylation of cysteines 3 and 4 that targets KillerRed to cellular membranes [Skene and Virág 1989]. pKillerRed-mem vector can be used as a source of MLS-KillerRed hybrid sequence. The vector backbone contains unique restriction sites that permit its excision and further insertion into expression vector of choice.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 E. coli, and f1 origin for single-stranded DNA production. SV40 polyadenylation signals (SV40 poly A) direct proper processing of the 3’-end of the reporter mRNA. 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 E. coli. Kanr /Neor gene is linked with herpes simplex virus (HSV) thymidine kinase (TK) polyadenylation signals.pKillerRed-mem vector can be transfected into mammalian cells by any known transfection method. CMV promoter provides strong, constitutive expression of memrane-targeted KillerRed in eukaryotic cells. If required, stable transformants can be selected using G418 [Gorman 1985].Suitable host strains for propagation in E. coli include DH5alpha, HB101, XL1-Blue, and other general purpose strains. Plasmid incompatibility group is pMB1/ColE1. The vector confers resistance to kanamycin (30 µg/ml) to E. coli hosts. Copy number in E. coli is about 500.

 

pKillerRed-Mem Sequence

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