CopyControl? cDNA, Gene & PCR Cloning Kit w/ Chem. Comp Cells

epicentre?CCPCR1CC?20 Rxns?CopyControl? cDNA

產品簡介：北京中北林格供應CopyControl? cDNA, Gene & PCR Cloning Kit w/ Chem. Comp Cells，中北林格是epicentre總代理，提供epicentre產品貨號報價查詢。其他包括lucigen總代理，cellscript總代理，Biosearch technologies總代理，illumina總代理，ICLLAB總代理，ImmunoReagents總代理。

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產品名稱:

CopyControl? cDNA, Gene & PCR Cloning Kit w/ Chem. Comp Cells

貨號:CCPCR1CC

規格:20 Rxns

存儲溫度:

Chemically Competent Cells are stored at -70℃, the remainder is stored at -20℃

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CopyControl? cDNA, Gene & PCR Cloning Kit w/ Chem. Comp Cells 詳細說明

epicentre?CCPCR1CC?20 Rxns?CopyControl? cDNA?詳細說明

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應用領域

• 有效克隆最多15 kb的任何插入片段，包括可能不穩定

  AIS107B

  或編碼對大腸桿菌宿主細胞有毒的蛋白質的那些片段。
• Efficient cloning of any insert up to 15 kb, including those that may be unstable or encode proteins that are toxic to the?E. coli?host cells.

如果cDNA克隆存在于宿主細胞中，每個細胞僅約一個拷貝（P. Carninci和W. Szybalski，個人交流），則可以從庫中獲得多達25％的cDNA克隆，這可能是由于有害作用降低所致對宿主細胞的代謝和表達的克隆序列及基因產物的毒性較低。CopyControl?cDNA，基因和PCR克隆試劑盒*提供了一種快速且改進的方法（圖1），用于克隆單拷貝數的任何平末端DNA片段（例如，基因組DNA，PCR產物或cDNA），并帶有具有根據需要誘導高拷貝數的能力（圖2）。試劑盒中提供的CopyControl pCC1?（Blunt）載體包含大腸桿菌?F因子單拷貝復制起點（ori?II）和誘導型高拷貝oriV.使用ori?V?不會發生復制，除非并且直到在TransforMax?EPI300?細胞的染色體（處于嚴格調控的啟動子的控制下）中誘導出trfA基因。因此，CopyControl cDNA，PCR或基因克隆可以單拷貝生長，以確保插入物穩定性和對宿主細胞有毒序列的成功克隆。然后，每個細胞最多可誘導200個拷貝，以用于所有下游應用，從而獲得高產量的DNA。

Up to ~25% more cDNA clones can be obtained in a library if the cDNA clones are present in the host cell at only about one copy per cell (P. Carninci and W. Szybalski, personal communication), presumably due to reduced detrimental effects on host cell metabolism and lower toxicity of expressed clone sequences and gene products. The CopyControl? cDNA, Gene, and PCR Cloning Kits* provide a rapid and improved method (Fig. 1) for cloning any blunt-end DNA fragments (e.g., genomic DNA, PCR products, or cDNA) at single-copy number, with the capability to induce to high-copy numbers as needed (Fig. 2). The CopyControl pCC1? (Blunt) Vector provided in the kits contains both the?E. coli?F-factor single-copy origin of replication (oriII) and an inducible high-copy?oriV. Replication does not occur using the?oriV unless and until the?trfA?gene in the chromosome of TransforMax? EPI300? cells (which is under the control of a tightly regulated promoter) is induced. Thus, CopyControl cDNA, PCR, or gene clones can be grown at single copy to ensure insert stability and successful cloning of sequences that are toxic to the host cell. Then, any desired clone can be induced up to 200 copies per cell for high yields of DNA for all downstream applications.

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好處

• 獲得更多的cDNA或PCR克隆和更完整的克隆文庫（表1）。
• 獲得較少的突變克隆。以高拷貝數選擇性地偏向對宿主細胞有毒性的基因中具有突變的克隆。
• 在1小時或更短的時間內篩選克隆的插入片段大小，而無需過夜培養或限制性內切酶消化（圖3）。
• Obtain more cDNA or PCR clones and more complete clone libraries (Table 1).
• Obtain fewer mutated clones. Clones having mutations in genes that are toxic to the host cell are selectively favored at high-copy number.
• Screen clones for insert size in 1 hour or less without the need for overnight cultures or restriction endonuclease digests (Fig. 3).

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CopyControl? cDNA, Gene & PCR Cloning Kit w/ Chem. Comp Cells?圖例說明

epicentre?CCPCR1CC?20 Rxns?CopyControl? cDNA?圖例說明

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圖1（單擊放大）。使用CopyControl?cDNA，基因和PCR克隆試劑盒，可在24小時內完成長達15 kb的插入片段的平末端平末端高效克隆。為了提高克隆的穩定性，每個細胞可以將克隆保持一個拷貝，然后，只要需要，任何克隆都可以被誘導為高拷貝數（每個細胞最多200個拷貝），從而獲得高產量的DNA。

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圖2.單拷貝CopyControl?克隆每個細胞可誘導200個拷貝。在1％瓊脂糖凝膠中通過電泳分析了來自相同數量的未誘導（-）細胞（泳道1）和未稀釋的誘導（+）細胞（泳道5）的DNA，該DNA包含10-kb CopyControl PCR克隆。通過目測比較稀釋的誘導樣品和未誘導樣品的染色強度來估計誘導細胞中的拷貝數。將來自誘導細胞的DNA以1：200（泳道2），1：100（泳道3）和1:50（泳道4）稀釋。

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圖3.從5-kb PCR產物中篩選克隆。使用CopyControl?cDNA，基因和PCR克隆試劑盒克隆PCR產物。使用試劑盒篩選了來自九個白色菌落的極小部分，確認了5kb插入片段的存在。M泳道，DNA大小階梯；泳道1，8 kb超螺旋DNA標記;?2-10道，隨機選擇的克隆。菌落篩選（包括凝膠電泳）的總時間約為60分鐘。

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貨號尺寸

CopyControl?cDNA，基因和PCR克隆試劑盒（帶電感受態細胞）???CCECPCR120個反應內容：?CopyControl?pCC1?（平直克隆就緒）載體，PCR沉淀溶液，PCR 10X克隆緩沖液，末端修復酶混合物，Fast-Link?DNA連接酶，EpiLyse?解決方案，EpiBlue?解決方案，CopyControl?Fosmid自動誘導解決方案，對照PCR產物，超螺旋DNA大小標記，無菌水，具有電抗性的TransforMax?EPI300??大腸桿菌。

CopyControl?cDNA，基因和PCR克隆試劑盒（具有化學感受態細胞）???CCPCR1CC20個反應內容：?CopyControl?pCC1?（平直克隆就緒）載體，PCR沉淀溶液，PCR 10X克隆緩沖液，末端修復酶混合物，Fast-Link?DNA連接酶，EpiLyse?解決方案，EpiBlue?解決方案，CopyControl?Fosmid自動誘導解決方案，對照PCR產物，超螺旋DNA大小標記，無菌水，具有化學活性的TransforMax?EPI300??大腸桿菌。

CopyControl?感應解決方案???CCIS12525毫升1,000X濃縮溶液。過濾器已消毒。

Catalog No.Size

CopyControl? cDNA, Gene and PCR Cloning Kit (with electrocompetent cells)???CCECPCR120 ReactionsContents:?CopyControl? pCC1? (Blunt Cloning-Ready) Vector, PCR Precipitation Solution, PCR 10X Cloning Buffer, End-Repair Enzyme Mix, Fast-Link? DNA Ligase, EpiLyse? Solution, EpiBlue? Solution, CopyControl? Fosmid Autoinduction Solution, Control PCR Product, Supercoiled DNA Size Marker, Sterile Water, Electrocompetent TransforMax? EPI300??E. coli.

CopyControl? cDNA, Gene and PCR Cloning Kit (with chemically competent cells)???CCPCR1CC20 ReactionsContents:?CopyControl? pCC1? (Blunt Cloning-Ready) Vector, PCR Precipitation Solution, PCR 10X Cloning Buffer, End-Repair Enzyme Mix, Fast-Link? DNA Ligase, EpiLyse? Solution, EpiBlue? Solution, CopyControl? Fosmid Autoinduction Solution, Control PCR Product, Supercoiled DNA Size Marker, Sterile Water, Chemically Competent TransforMax? EPI300??E. coli.

CopyControl? Induction Solution???CCIS12525 ml1,000X concentrated solution. Filter sterilized.

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