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BASIC:一种简单且精确的模块化DNA组装方法。

BASIC: A Simple and Accurate Modular DNA Assembly Method.

作者信息

Storch Marko, Casini Arturo, Mackrow Ben, Ellis Tom, Baldwin Geoff S

机构信息

Department of Life Sciences, Centre for Synthetic Biology and Innovation, Imperial College London, London, SW7 2AZ, UK.

Department of Bioengineering, Centre for Synthetic Biology and Innovation, Imperial College London, London, SW7 2AZ, UK.

出版信息

Methods Mol Biol. 2017;1472:79-91. doi: 10.1007/978-1-4939-6343-0_6.

Abstract

Biopart Assembly Standard for Idempotent Cloning (BASIC) is a simple, accurate, and robust DNA assembly method. The method is based on linker-mediated DNA assembly and provides highly accurate DNA assembly with 99 % correct assemblies for four parts and 90 % correct assemblies for seven parts [1]. The BASIC standard defines a single entry vector for all parts flanked by the same prefix and suffix sequences and its idempotent nature means that the assembled construct is returned in the same format. Once a part has been adapted into the BASIC format it can be placed at any position within a BASIC assembly without the need for reformatting. This allows laboratories to grow comprehensive and universal part libraries and to share them efficiently. The modularity within the BASIC framework is further extended by the possibility of encoding ribosomal binding sites (RBS) and peptide linker sequences directly on the linkers used for assembly. This makes BASIC a highly versatile library construction method for combinatorial part assembly including the construction of promoter, RBS, gene variant, and protein-tag libraries. In comparison with other DNA assembly standards and methods, BASIC offers a simple robust protocol; it relies on a single entry vector, provides for easy hierarchical assembly, and is highly accurate for up to seven parts per assembly round [2].

摘要

幂等克隆生物部件组装标准(BASIC)是一种简单、准确且稳健的DNA组装方法。该方法基于接头介导的DNA组装,对于四个部件的组装,能提供99%的正确组装率,对于七个部件的组装,正确组装率为90%,从而实现高度准确的DNA组装[1]。BASIC标准为所有部件定义了一个单一的入口载体,其两侧为相同的前缀和后缀序列,并且其幂等性质意味着组装后的构建体以相同格式返回。一旦一个部件被适配为BASIC格式,它就可以放置在BASIC组装中的任何位置,而无需重新格式化。这使得实验室能够构建全面且通用的部件库,并有效地共享它们。通过直接在用于组装的接头上编码核糖体结合位点(RBS)和肽接头序列的可能性,BASIC框架内的模块化得到了进一步扩展。这使得BASIC成为一种用于组合部件组装的高度通用的文库构建方法,包括启动子、RBS、基因变体和蛋白质标签文库的构建。与其他DNA组装标准和方法相比,BASIC提供了一个简单稳健的方案;它依赖于单一的入口载体,便于进行分层组装,并且在每个组装轮次中多达七个部件时都具有高度准确性[2]。

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