pGL4.43载体基本信息

　　出品公司:Promega

　　载体名称:pGL4.43 ; pGL4.43[luc2P/XRE/Hygro]

　　质粒类型:信号通路报告载体;哺乳动物载体;萤火虫荧光素酶报告载体

　　高拷贝/低拷贝:--

　　克隆方法:限制性内切酶，多克隆位点

　　启动子:Minimal Promotor

　　载体大小:6067 bp

　　5' 测序引物及序列:--

　　3' 测序引物及序列:--

　　载体标签:luc2P

　　载体抗性:氨苄青霉素

　　筛选标记:潮霉素(Hygromycin)

　　克隆菌株:TOP10等常规菌株

　　宿主细胞(系):HEK293等

　　备注:pGL4.43[luc2P/XRE/Hygro]载体是信号通路报告载体;含异己物应答元件XRE;含萤火虫荧光素酶报告基因luc2P。

　　产品目录号:E3751

　　稳定性:稳表达

　　组成型/诱导型:组成型

　　病毒/非病毒:非病毒

　　pGL4.43载体质粒图谱和多克隆位点信息

　　pGL4.43载体简介

　　The pGL4.43[luc2P/XRE/Hygro] Vector contains three copies of a xenobiotic response element (XRE) that drives transcription of the luciferase reporter gene luc2P (Photinus pyralis). luc2P is a synthetically-derived luciferase sequence with humanized codon optimization that is designed for high expression and reduced anomalous transcription. The luc2P gene contains hPEST, a protein destabilization sequence, which allows luc2P protein levels to respond more quickly than those of luc2 to induction of transcription. The vector backbone contains an ampicillin resistance gene to allow selection in E. coli and a gene for hygromycin resistance to allow selection of stably transfected mammalian cell lines.

　　Example Protocol

　　In this example protocol, the pGL4.43[luc2P/XRE/Hygro] Vector is used to measure

　　activation of the XRE in HepG2 cells upon treatment with TCDD. The pGL4.75 Vector

　　(encoding Renilla luciferase) is used as a normalization control. In designing such

　　experiments, it is important that the chosen cell type can be transfected efficiently and

　　that it expresses the proper components of the signaling pathway of interest in order to

　　generate the biological response. Protocol optimization may be required for your

　　particular cell type and assay conditions.

　　实验材料

　　DMEM (Life Technologies Cat.# 11995)

　　Complete medium [DMEM supplemented with 10% fetal bovine serum (DMEM/FBS;

　　Life Technologies Cat.# 16000] and 1X NEAA [Life Technologies Cat.# 11140])

　　Dulbecco’s PBS (DPBS; Life Technologies Cat. # 14190)

　　0.05% Tryspin-EDTA (Life Technologies Cat.# 25300)

　　Charcoal-stripped FBS (Life Technologies Cat.# 126776-011)

　　Opti-MEM I (Life Technologies Cat.# 31985)

　　FuGENE HD Transfection Reagent (Cat.# E2311)

　　TCDD (2.3.7.8-Tetrachlorodibenzo-p-dioxin; AccuStandard Cat.# D-404N)

　　DMSO (Sigma Cat.# D2650)

　　Dual-Glo Luciferase Assay System (Cat.# E2940)

　　HepG2 cells

　　pGL4.75[hRluc/CMV] Vector (Cat.# E6931)

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