A Modular Assembly Platform for Rapid Generation of DNA Constructs
A Modular Assembly Platform for Rapid Generation of DNA Constructs
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DOI：
10.1038/srep16836
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发表时间：
2016-02-18
影响因子：
4.6
通讯作者：
Jacks, Tyler
中科院分区：
文献类型：
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作者：
Akama-Garren, Elliot H.;Joshi, Nikhil S.;Jacks, Tyler
传统的克隆方法在可同时操作的DNA片段数量上存在局限性，这极大地减缓了分子组装的速度。在此我们描述了GMAP，一个基于吉布森组装的模块化组装平台，它由一系列启动子和基因组成，允许一步生产DNA构建体。GMAP利用模块化基因组件促进表达和病毒构建体的快速组装，以及利用与环境相关的基因组元件促进日益复杂的基因工具的组装。我们的数据证明了GMAP在合成启动子验证、有效RNAi构建体鉴定、诱导型慢病毒系统建立、基因工程小鼠模型中的肿瘤起始以及产生敲入小鼠的基因靶向方面的适用性。GMAP代表了一种重组DNA技术，旨在广泛传播并易于适应其他用途，例如合成生物学、基因筛选和CRISPR - Cas9。
Traditional cloning methods have limitations on the number of DNA fragments that can be simultaneously manipulated, which dramatically slows the pace of molecular assembly. Here we describe GMAP, a Gibson assembly-based modular assembly platform consisting of a collection of promoters and genes, which allows for one-step production of DNA constructs. GMAP facilitates rapid assembly of expression and viral constructs using modular genetic components, as well as increasingly complicated genetic tools using contextually relevant genomic elements. Our data demonstrate the applicability of GMAP toward the validation of synthetic promoters, identification of potent RNAi constructs, establishment of inducible lentiviral systems, tumor initiation in genetically engineered mouse models, and gene-targeting for the generation of knock-in mice. GMAP represents a recombinant DNA technology designed for widespread circulation and easy adaptation for other uses, such as synthetic biology, genetic screens, and CRISPR-Cas9.