In-Fusion BioBrick assembly and re-engineering.
In-Fusion BioBrick assembly and re-engineering.
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DOI：
10.1093/nar/gkq179
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发表时间：
2010-05
影响因子：
14.9
通讯作者：
Sauro HM
中科院分区：
文献类型：
--
作者：
Sleight SC;Bartley BA;Lieviant JA;Sauro HM
遗传电路可以由称为 BioBricks 的标准化生物部件组装而成。 BioBrick 的例子包括启动子、核糖体结合位点、编码序列和转录终止子。标准 BioBrick 组装通常涉及限制性酶消化和一次连接两个 BioBrick。这里描述的方法是一种替代组装策略，允许使用 Clontech In-Fusion PCR 克隆试剂盒快速组装和重新设计两个或多个 PCR 扩增的 BioBrick。该方法允许执行大量并行组装，并且是混合和匹配 BioBricks 的灵活方式。 In-Fusion 组装可以通过使用简单的引物设计规则实现半标准化，从而最大限度地减少规划组装反应所需的时间。我们描述了使用各种同源性和引物长度的 In-Fusion 组装遗传电路的成功率和突变率。我们还通过 BioBrick 组装和重新设计的六个具体示例展示了该方法的成功和灵活性。这些示例包括两个基本零件的装配、零件交换、删除、插入和三向 In-Fusion 装配。
Genetic circuits can be assembled from standardized biological parts called BioBricks. Examples of BioBricks include promoters, ribosome-binding sites, coding sequences and transcriptional terminators. Standard BioBrick assembly normally involves restriction enzyme digestion and ligation of two BioBricks at a time. The method described here is an alternative assembly strategy that allows for two or more PCR-amplified BioBricks to be quickly assembled and re-engineered using the Clontech In-Fusion PCR Cloning Kit. This method allows for a large number of parallel assemblies to be performed and is a flexible way to mix and match BioBricks. In-Fusion assembly can be semi-standardized by the use of simple primer design rules that minimize the time involved in planning assembly reactions. We describe the success rate and mutation rate of In-Fusion assembled genetic circuits using various homology and primer lengths. We also demonstrate the success and flexibility of this method with six specific examples of BioBrick assembly and re-engineering. These examples include assembly of two basic parts, part swapping, a deletion, an insertion, and three-way In-Fusion assemblies.
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影响因子：
14.9
作者：
Gibson DG
通讯作者：
Gibson DG
用户融合：一种快速有效的方法，用于同时融合和克隆多个PCR产品。
影响因子：
14.9
作者：
Geu-Flores, Fernando;Nour-Eldin, Hussam H.;Nielsen, Morten T.;Halkier, Barbara A.
通讯作者：
Halkier, Barbara A.
影响因子：
48
作者：
Gibson, Daniel G.;Young, Lei;Smith, Hamilton O.
通讯作者：
Smith, Hamilton O.
影响因子：
46.9
作者：
Canton, Barry;Labno, Anna;Endy, Drew
通讯作者：
Endy, Drew
影响因子：
5.6
作者：
Anderson JC;Dueber JE;Leguia M;Wu GC;Goler JA;Arkin AP;Keasling JD
通讯作者：
Keasling JD