Biofoundry-Scale DNA Assembly Validation Using Cost-Effective High-Throughput Long-Read Sequencing
Biofoundry-Scale DNA Assembly Validation Using Cost-Effective High-Throughput Long-Read Sequencing
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DOI：
10.1021/acssynbio.3c00589
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发表时间：
2024-02-08
影响因子：
4.7
通讯作者：
Fragkoudis，Rennos
中科院分区：
文献类型：
--
作者：
Vegh，Peter;Donovan，Sophie;Fragkoudis，Rennos
生物工厂是自动化的高通量设施，专门从事设计，构建和测试工程/合成DNA构建体（质粒），通常来自遗传部分。该过程的关键步骤是评估组装的DNA构建体对所需设计的保真度。目前用于此目的的方法是限制性消化或PCR，然后进行片段分析和测序。Edinburgh Genome Foundry（EGF）最近使用Oxford Nanopore测序技术建立了单分子测序质量控制步骤，沿着配套Nextflow管道和Python包，以进行深入分析并生成详细报告。我们的软件使研究质粒的研究人员，包括生物基金科学家，能够快速分析和解释测序数据。总之，我们创建了一个实验室和软件协议，使用Nanopore长读段验证组装、克隆或编辑的质粒，这可以作为遗传学、合成生物学和测序界的有用资源。
Biofoundries are automated high-throughput facilities specializing in the design, construction, and testing of engineered/synthetic DNA constructs (plasmids), often from genetic parts. A critical step of this process is assessing the fidelity of the assembled DNA construct to the desired design. Current methods utilized for this purpose are restriction digest or PCR followed by fragment analysis and sequencing. The Edinburgh Genome Foundry (EGF) has recently established a single-molecule sequencing quality control step using the Oxford Nanopore sequencing technology, along with a companion Nextflow pipeline and a Python package, to perform in-depth analysis and generate a detailed report. Our software enables researchers working with plasmids, including biofoundry scientists, to rapidly analyze and interpret sequencing data. In conclusion, we have created a laboratory and software protocol that validates assembled, cloned, or edited plasmids, using Nanopore long-reads, which can serve as a useful resource for the genetics, synthetic biology, and sequencing communities.