DNA assembly for synthetic biology: from parts to pathways and beyond
DNA assembly for synthetic biology: from parts to pathways and beyond
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DOI：
10.1039/c0ib00070a
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发表时间：
2011-01-01
影响因子：
2.5
通讯作者：
Baldwin, Geoff S.
中科院分区：
文献类型：
--
作者：
Ellis, Tom;Adie, Tom;Baldwin, Geoff S.
将DNA从小片段组装成大结构的技术最近取得了重大发展，成为实现合成生物学愿景的关键技术。随着全基因合成的成本不断降低，另一端的全基因组合成已将我们的视野扩展到完全工程化的合成细胞的前景。然而，最近被证明的合成基因组规模 DNA 的能力与我们合理设计生物设备的能力相矛盾，后者明显落后。合成生物学中的大多数工作都是在中等规模上进行的，通过组合构建来自模块化生物部件组件的网络和代谢途径。实现工程生物电路的快速原型制作需要根据特定的架构进行快速、可靠的 DNA 组装。显然，DNA 组装现在是合成生物学发展的一项限制技术。目前的技术采用标准化限制性酶组装方案，例如BioBricks (TM)、BglBricks和Golden Gate方法。另外，序列无关的重叠技术，例如 In-Fusion (TM)、SLIC 和 Gibson 等温组装，对于较大的组装越来越流行，并且酵母和芽孢杆菌中的体内 DNA 组装似乎适合染色体制造。重要的是要考虑不同技术的使用如何影响构造的结果，因为组装技术可以指导可制造的系统的架构和多样性。这篇综述对最近的 DNA 组装策略进行了严格的审查，并考虑了合成生物学这一重要的促进方面在未来如何发展。
The assembly of DNA from small fragments into large constructs has seen significant recent development, becoming a pivotal technology in the ability to implement the vision of synthetic biology. As the cost of whole gene synthesis is decreasing, whole genome synthesis at the other end of the spectrum has expanded our horizons to the prospect of fully engineered synthetic cells. However, the recently proven ability to synthesise genome-scale DNA is at odds with our ability to rationally engineer biological devices, which lags significantly behind. Most work in synthetic biology takes place on an intermediate scale with the combinatorial construction of networks and metabolic pathways from registries of modular biopart components. Implementation for rapid prototyping of engineered biological circuits requires quick and reliable DNA assembly according to specific architectures. It is apparent that DNA assembly is now a limiting technology in advancing synthetic biology. Current techniques employ standardised restriction enzyme assembly protocols such as BioBricks (TM), BglBricks and Golden Gate methods. Alternatively, sequence-independent overlap techniques, such as In-Fusion (TM), SLIC and Gibson isothermal assembly are becoming popular for larger assemblies, and in vivo DNA assembly in yeast and bacillus appears adept for chromosome fabrication. It is important to consider how the use of different technologies may impact the outcome of a construction, since the assembly technique can direct the architecture and diversity of systems that can be made. This review provides a critical examination of recent DNA assembly strategies and considers how this important facilitating aspect of synthetic biology may proceed in the future.