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lucigen 49003-1 Expresso? T7 SUMO Cloning

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lucigen 總代理

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主要規(guī)格：
5 rxns

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科研

lucigen 49003-1 Expresso? T7 SUMO Cloning 產(chǎn)品簡介：北京中北林格供應(yīng)lucigen 49003-1 Expresso? T7 SUMO Cloning，中北林格是epicentre總代理，提供epicentre產(chǎn)品貨號報價查詢。其他包括lucigen總代理，cellscript總代理，Biosearch technologies總代理，epicentre總代理，illumina總代理，ICLLAB總代理，ImmunoReagents總代理。
產(chǎn)品名稱:Expresso? T7 SUMO Cloning and Expression System 貨號:49003-1 規(guī)格:5 rxns 存儲溫度:-20o & -80o C 品牌：lucigen / epicentre
供應(yīng)商：中北林格
產(chǎn)地：us
發(fā)貨地：北京
lucigen 49003-1 Expresso? T7 SUMO Cloning 產(chǎn)品說明書產(chǎn)品描述可以快速，容易的克隆和表達(dá)困難蛋白。是基于定向PCR克隆系統(tǒng)所研制的克隆表達(dá)系統(tǒng)。 Expresso saves you a day! 完整的克隆表達(dá)系統(tǒng) Expresso? T7 克隆和蛋白表達(dá)系統(tǒng)是研制出的一款快速，簡單，有效的定向克隆和表達(dá)PCR擴(kuò)增產(chǎn)物的系統(tǒng)。系統(tǒng)含有預(yù)處理的pETite? N或C端組氨酸標(biāo)簽克隆載體，兩個感受態(tài)細(xì)胞，存在于一個單獨(dú)的轉(zhuǎn)化瓶中。高效HI-Control? 10G化學(xué)感受態(tài)細(xì)胞能穩(wěn)定克隆，HI-Control BL21(DE3)感受態(tài)細(xì)胞提供了嚴(yán)謹(jǐn)?shù)目煽氐鞍妆磉_(dá)。因此可以避免泄露型T7啟動子系統(tǒng)存在的表達(dá)問題。N或C端的組氨酸標(biāo)簽蛋白可以使你快速的使用鎳柱純化表達(dá)蛋白。系統(tǒng)還具有在N端編碼6xHis SUMO融合標(biāo)簽可以改善表達(dá)蛋白的可溶性。其他表達(dá)系統(tǒng)為鼠李糖啟動子控制的表達(dá)。 5秒定向克隆PCR擴(kuò)增基因 Expresso? T7 克隆和蛋白表達(dá)系統(tǒng)使用Expressioneering ?技術(shù)，是一個無酶的重組克隆策略到無縫基因載體整合過程。使用帶有18bp載體互補(bǔ)序列的引物對靶向基因進(jìn)行PCR擴(kuò)增。不同于其他的限制酶方法或無連接酶方法，不需要進(jìn)一步純化和酶處理PCR產(chǎn)物。只需要簡單混合1μl不需純化的PCR反應(yīng)產(chǎn)物至預(yù)處理的pETite? T7表達(dá)載體，立即轉(zhuǎn)化至HI-Control? 10G化學(xué)感受態(tài)細(xì)胞（如圖1）。優(yōu)點(diǎn)：省時，無酶的克隆體系，利用克隆載體和細(xì)胞只需數(shù)秒完成。高效：?90%的重組子。嚴(yán)謹(jǐn)控制表達(dá)，N端或C端帶有6組氨酸標(biāo)簽蛋白。 Expressioneering? 技術(shù)，所有Expresso?產(chǎn)品的特征為無連接酶克隆。克隆載體構(gòu)建，使研究時間節(jié)省數(shù)十小時全新New pETite T7 載體包含: 用于高表達(dá)的T7強(qiáng)啟動子用于蛋白快速純化的位于N端或C端的組氨酸蛋白融合標(biāo)簽。大小只有2.2kb，易于下游操作。專利克隆技術(shù)增加了克隆效率。 lucigen 49003-1 Expresso? T7 SUMO Cloning 圖例說明 pETite vector maps Figure 2. pETite T7 expression vectors: Small size (2.2 kb vs. 5.4 kb for pET) for easier manipulation, including targeted mutagenesis. Vectors are pre-processed for instant enzyme-free cloning of PCR products with a choice of amino-terminal or carboxyl-terminal fusion of 6xHis tag to protein of interest. 使用HI-Control 10G 化學(xué)感受態(tài)細(xì)胞獲得高效率克隆高效的HI-Control 10G 化學(xué)感受態(tài)細(xì)胞可以確保篩選得到插入正確的克隆。對于大多數(shù)基因，> 90%的克隆都是靶向基因插入方向正確的陽性克隆（如圖三）。 High cloning efficiency with Expresso T7 System Figure 3. High cloning efficiency with Expresso T7 System. Pre-processed pETite C-His vector was mixed with 1 μl of unpurified PCR gene product and transformed into HI-Control 10G Chemically Competent Cells. Colony PCR was performed on 18 randomly chosen colonies; 17 of 18 contained insert of the correct size. HI-Control BL21(DE3)細(xì)胞控制泄漏蛋白表達(dá) HI-Control BL21(DE3) 細(xì)胞含有高水平lac抑制物，可以保證嚴(yán)謹(jǐn)控制T7 RNA聚合酶的表達(dá)。嚴(yán)格控制意味著更好的適應(yīng)潛在毒性基因產(chǎn)物（圖4）。 Expresso vs. pET28a Figure 4. Expression of various proteins using the Expresso T7 System versus a pET Vector. Genes encoding a DNA polymerase, a blue fluorescent protein (BFP), or ATP synthase b subunit (membrane protein) were cloned into pET28a or pETite vectors with N-terminal or C-terminal 6xHis tags (as indicated). pET28a clones were transformed into standard BL21(DE3) cells, and pETite clones were transformed into HI-Control BL21(DE3) cells for expression. Cultures were grown in LB at 37°C to an OD600 of 0.5 to 0.7 (odd-numbered lanes) and induced for 3 hours with 1 mM IPTG (even-numbered lanes). Cells were pelleted and lysed directly in SDS-PAGE loading buffer, and 0.05 OD equivalents were analyzed by gel electrophoresis. The gel was stained with Coomassie blue. Expression and purification of active soluble fluorescent protein. Figure 5. Purification of a 6xHis tagged fluorescent protein. HI-Control BL21(DE3) cells harboring pETite C-His vector containing a yellow fluorescent protein (YFP) gene were grown at 37°C in LB media to an OD600 of 0.6 (lane 1), then induced with 1 mM IPTG for 4 hours (lane 2). Cells were harvested and lysed by sonication in 300 mM NaCl, 50 mM Tris-HCl pH 8.0. Cleared lysate was loaded onto an Ni-NTA Sepharose? Column. Column flow-through (lane 3, FT) and wash (lane 4, W) fractions were collected. The bound YFP, shown in the column at left, was eluted with wash buffer containing 300 mM imidazole (lanes 5-12, E1-E8). Fluorescent fractions align with the pure protein as shown on the gel. lucigen 49003-1 Expresso? T7 SUMO Cloning 貨號詳情 Contact your local distributor for pricing Product Description Size Cat. No. Expresso? T7 Cloning and Expression System, N-His 5 rxns 49001-1 10 rxns 49001-2 Expresso? T7 Cloning and Expression System, C-His 5 rxns 49002-1 10 rxns 49002-2 Expresso? T7 Cloning and Expression System, N/C-His Combo, 5 of each of N-His and C-His 10 rxns 49000-1 Expresso? T7 SUMO Cloning and Expression System 5 rxns 49003-1 10 rxns 49003-2 HI-Control? 10G Chemically Competent Cells (SOLOs) 12 rxns (SOLOs) 60110-1 HI-Control? BL21(DE3) Chemically Competent Cells (SOLOs) 12 rxns (SOLOs) 60435-1 SUMO Express Protease 200 U 30801-2 ORDER INFORMATION T7克隆和表達(dá)系統(tǒng)含有預(yù)處理的pETite? N-His and/or pETite C-His 載體DNA, 用于克隆的HI-Control? 10G化學(xué)感受態(tài)細(xì)胞和用于蛋白表達(dá)的HI-Control BL21(DE3) 化學(xué)感受態(tài)細(xì)胞。同時還包括N端或C端組氨酸標(biāo)簽陽性對照插入DNA和轉(zhuǎn)化陽性對照PUC DNA . T7 SUMO克隆和表達(dá)系統(tǒng)含有預(yù)處理的 pETite? N-His SUMO 載體 DNA, 用于克隆的HI-Control? 10G 化學(xué)感受態(tài)細(xì)胞和用于蛋白表達(dá)的 HI Control BL21(DE3) 化學(xué)感受態(tài)細(xì)胞，也包括SUMO 陽性對照插入DNA, 轉(zhuǎn)化陽性對照 pUC DNA, SUMO表達(dá)蛋白酶, SUMO 切割控制蛋白, 鑒定克隆的正反向PCR引物。對此產(chǎn)品有興趣，可聯(lián)系中北林格獲取更多詳細(xì)信息。

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