As a biorefinery platform host, Escherichia coli has been used extensively to produce metabolites of commercial interest. Integration of foreign DNA onto the bacterial genome allows for stable expression overcoming the need for plasmid expression and its associated instability. Despite the development of numerous tools and genome editing technologies, the question of where to incorporate a synthetic pathway remains unanswered. To address this issue, we studied the genomic expression in E. coli and linked it not only to 26 rationally selected genomic locations, but also to the gene direction in relation to the DNA replication fork, to the carbon and nitrogen source, to DNA folding and supercoiling, and to metabolic burden. To enable these experiments, we have designed a fluorescent expression cassette to eliminate specific local effects on gene expression. Overall it can be concluded that although the expression range obtained by changing the genomic location of a pathway is small compared to the range typically seen in promoter-RBS libraries, the effect of culture medium, environmental stress and metabolic burden can be substantial. The characterization of multiple effects on genomic expression, and the associated libraries of well-characterized strains, will only stimulate and improve the creation of stable production hosts fit for industrial settings.

作为生物精炼平台宿主，大肠杆菌已被广泛用于生产具有商业价值的代谢物。将外源 DNA 整合到细菌基因组上可以实现稳定表达，克服了对质粒表达的需要及其相关的不稳定性。尽管开发了许多工具和基因组编辑技术，但在何处整合合成途径的问题仍未得到解答。为了解决这个问题，我们研究了大肠杆菌中的基因组表达并将其不仅与 26 个合理选择的基因组位置相关联，还与 DNA 复制叉、碳和氮源、DNA 折叠和超螺旋以及代谢负担相关的基因方向相关联。为了进行这些实验，我们设计了一个荧光表达盒来消除对基因表达的特定局部影响。总的来说，可以得出结论，虽然通过改变通路的基因组位置获得的表达范围与启动子-RBS 文库中常见的范围相比较小，但培养基、环境压力和代谢负担的影响可能很大。对基因组表达的多重影响的表征，以及表征良好的菌株的相关文库，