Plasmid DNA is a vital biological tool for molecular cloning and transgene expression of recombinant proteins; however, decades ago, it has become an exceptionally appealing as a potential biopharmaceutical product as genetic immunization for animal and human use. The demand for large-quantity production of DNA vaccines also increases. Thus, we, herein, presented a systematic approach for process characterization of fed-batch Escherichia coli DH5α fermentation producing a porcine DNA vaccine. Design of Experiments (DoE) was employed to determine process parameters that have impacts on a critical quality attribute of the product, which is the active form of plasmid DNA referred as supercoiled plasmid DNA content, as well as the performance attributes, which are volumetric yield and specific yield from fermentation. The parameters of interest were temperature, pH, dissolved oxygen, cultivation time, and feed rate. Using the definitive-screening design, there were 16 runs, including 3 additional center points to create the predictive model, which then was used to simulate the operational ranges for capability analysis.

中文翻译：

---

DNA 疫苗生产的确定性筛选设计方法的工艺表征


质粒DNA是重组蛋白分子克隆和转基因表达的重要生物学工具；然而，几十年前，它已成为一种极具吸引力的潜在生物制药产品，可用于动物和人类的基因免疫。 DNA疫苗大批量生产的需求也随之增加。因此，我们在此提出了一种用于分批补料大肠杆菌 DH5α 发酵生产猪 DNA 疫苗的过程表征的系统方法。采用实验设计 (DoE) 来确定对产品关键质量属性（即质粒 DNA 的活性形式，称为超螺旋质粒 DNA 含量）以及性能属性（体积产量）有影响的工艺参数和发酵比产率。感兴趣的参数是温度、pH、溶解氧、培养时间和进料速率。使用最终筛选设计，进行了 16 次运行，包括 3 个额外的中心点来创建预测模型，然后使用该模型来模拟能力分析的操作范围。