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pGL4.43載體基本信息

　　出品公司:Promega

　　載體名稱:pGL4.43 ; pGL4.43[luc2P/XRE/Hygro]

　　質(zhì)粒類型:信號(hào)通路報(bào)告載體;哺乳動(dòng)物載體;螢火蟲熒光素酶報(bào)告載體

　　高拷貝/低拷貝:--

　　克隆方法:限制性內(nèi)切酶，多克隆位點(diǎn)

　　啟動(dòng)子:Minimal Promotor

　　載體大小:6067 bp

　　5' 測(cè)序引物及序列:--

　　3' 測(cè)序引物及序列:--

　　載體標(biāo)簽:luc2P

　　載體抗性:氨芐青霉素

　　篩選標(biāo)記:潮霉素(Hygromycin)

　　克隆菌株:TOP10等常規(guī)菌株

　　宿主細(xì)胞(系):HEK293等

　　備注:pGL4.43[luc2P/XRE/Hygro]載體是信號(hào)通路報(bào)告載體;含異己物應(yīng)答元件XRE;含螢火蟲熒光素酶報(bào)告基因luc2P。

　　產(chǎn)品目錄號(hào):E3751

　　穩(wěn)定性:穩(wěn)表達(dá)

　　組成型/誘導(dǎo)型:組成型

　　病毒/非病毒:非病毒

　　pGL4.43載體質(zhì)粒圖譜和多克隆位點(diǎn)信息

　　pGL4.43載體簡(jiǎn)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.

　　實(shí)驗(yàn)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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