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Ribosomal binding site switching: an effective strategy for high-throughput cloning constructions. PLoS One. 2012 誰幫忙在web of science 上幫忙查詢下 |
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liouzhan654
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感謝參與，應(yīng)助指數(shù) +1
guangyi: 金幣+5, 謝謝 2016-02-21 11:14:47
感謝參與，應(yīng)助指數(shù) +1
guangyi: 金幣+5, 謝謝 2016-02-21 11:14:47
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目前被引用次數(shù)為0次 Ribosomal Binding Site Switching: An Effective Strategy for High-Throughput Cloning Constructions 作者:Hu, YB (Hu, Yangbo)[ 1 ] ; Feng, LP (Feng, Lipeng)[ 1,2 ] ; Li, YL (Li, Yunlong)[ 1,2 ] ; Zhang, Y (Zhang, Yong)[ 1 ] ; Lu, P (Lu, Pei)[ 1 ] ; Rayner, S (Rayner, Simon)[ 1 ] ; Chen, SY (Chen, Shiyun)[ 1 ] 查看 ResearcherID 和 ORCID PLOS ONE 卷: 7 期: 11 文獻(xiàn)號(hào): e50142 DOI: 10.1371/journal.pone.0050142 出版年: NOV 21 2012 查看期刊信息 摘要 Direct cloning of PCR fragments by TA cloning or blunt end ligation are two simple methods which would greatly benefit high-throughput (HTP) cloning constructions if the efficiency can be improved. In this study, we have developed a ribosomal binding site (RBS) switching strategy for direct cloning of PCR fragments. RBS is an A/G rich region upstream of the translational start codon and is essential for gene expression. Change from A/G to T/C in the RBS blocks its activity and thereby abolishes gene expression. Based on this property, we introduced an inactive RBS upstream of a selectable marker gene, and designed a fragment insertion site within this inactive RBS. Forward and reverse insertions of specifically tailed fragments will respectively form an active and inactive RBS, thus all background from vector self-ligation and fragment reverse insertions will be eliminated due to the non-expression of the marker gene. The effectiveness of our strategy for TA cloning and blunt end ligation are confirmed. Application of this strategy to gene over-expression, a bacterial two-hybrid system, a bacterial one-hybrid system, and promoter bank construction are also verified. The advantages of this simple procedure, together with its low cost and high efficiency, makes our strategy extremely useful in HTP cloning constructions. 關(guān)鍵詞 KeyWords Plus  OSITIVE-SELECTION VECTORS; PCR PRODUCTS; ESCHERICHIA-COLI; BACILLUS-SUBTILIS; IN-VIVO; GENE; RECOMBINATION; SEQUENCE作者信息 通訊作者地址: Chen, SY (通訊作者) 顯示增強(qiáng)組織信息的名稱 Chinese Acad Sci, Wuhan Inst Virol, Key Lab Agr & Environm Microbiol, Wuhan, Peoples R China. 地址: 顯示增強(qiáng)組織信息的名稱 [ 1 ] Chinese Acad Sci, Wuhan Inst Virol, Key Lab Agr & Environm Microbiol, Wuhan, Peoples R China 顯示增強(qiáng)組織信息的名稱 [ 2 ] Chinese Acad Sci, Grad Univ, Beijing, Peoples R China 電子郵件地址:sychen@wh.iov.cn 基金資助致謝 基金資助機(jī)構(gòu) 授權(quán)號(hào) National Natural Science Foundation of China 31170133 National Transgenic Program of China 2011ZX08009-003-001-003 查看基金資助信息 出版商 PUBLIC LIBRARY SCIENCE, 1160 BATTERY STREET, STE 100, SAN FRANCISCO, CA 94111 USA 類別 / 分類 研究方向:Science & Technology - Other Topics Web of Science 類別:Multidisciplinary Sciences 文獻(xiàn)信息 文獻(xiàn)類型:Article 語種:English 入藏號(hào): WOS:000311821000189 PubMed ID: 23185557 ISSN: 1932-6203 期刊信息 Impact Factor (影響因子): Journal Citation Reports® 其他信息 IDS 號(hào): 047ER Web of Science 核心合集中的 "引用的參考文獻(xiàn)": 30 Web of Science 核心合集中的 "被引頻次": 0 |
2樓2016-02-21 11:11:37
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3樓2016-02-21 11:48:27
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