■ 基本信息
啟動(dòng)子: | CMV |
復(fù)制子: | pUC |
終止子: | SV40 poly(A) signal |
質(zhì)粒分類: | 哺乳細(xì)胞質(zhì)粒；哺乳報(bào)告質(zhì)粒；熒光定位質(zhì)粒 |
質(zhì)粒大小: | 4787bp |
質(zhì)粒標(biāo)簽: | C-mem，C-KillerRed |
原核抗性: | Kan |
真核抗性: | G418 |
克隆菌株: | DH5a |
培養(yǎng)條件: | 37度 |
表達(dá)宿主: | 293T等哺乳細(xì)胞 |
備注: | 哺乳細(xì)胞紅色熒光膜定位質(zhì)粒 |
■ 質(zhì)粒屬性
質(zhì)粒宿主: | 哺乳細(xì)胞 |
質(zhì)粒用途: | 定位 |
片段類型: | ORF |
片段物種: | 空載體 |
原核抗性: | Kan |
真核抗性: | G418
|
熒光標(biāo)記: | 紅色
|
■ 質(zhì)粒簡介
pKillerRed-mem質(zhì)粒是一個(gè)哺乳細(xì)胞紅色熒光定位于細(xì)胞膜的質(zhì)粒。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.
■ 質(zhì)粒圖譜
■ 質(zhì)粒序列
質(zhì)粒序列請下載：
ZK531pKillerRed-Mem細(xì)胞膜定位質(zhì)粒.txt
