Manufacturing large plasmid DNA - new approaches

在经历了一段相对平静的时期后，人们对基于质粒dna的疫苗和疗法的兴趣重新燃起。尽管在过去二十年中产生了大量关于质粒加工的数据，但在扩大质粒DNA的生产和纯化时仍然存在挑战，特别是如果现在预测的高需求成为现实，特别是疫苗接种。这些范围从低生产力的微生物发酵下游加工困难和缺乏一个平台的过程。目前的研究建立在伦敦大学学院的大量出版物和专业知识的基础上，以解决质粒DNA可制造性的一些瓶颈。假设：紧密形成的超卷曲DNA可以潜在地克服大规模生产的大型超卷曲质粒数量和质量低的挑战。这项研究的重点是我们实验室最近的研究，我们已经成功地证明了我们可以制造出尺寸缩小的紧密超螺旋DNA。这已经在6.5 Kb的质粒中实现了，我们将把它扩展到更大的质粒（超过20Kb），在那里可以包装更多的信息。然后，我们将研究在发酵罐生产的挑战，随后是回收和初始分离步骤。

There has been a resurgence of interest in plasmid DNA-based vaccines and therapeutics after a relatively quiet period. In spite of large volume of data generated over the past two decades on plasmid processing, there are still challenges that persist when scaling up the production and purification of plasmid DNA, especially if the now projected high demand materializes specially for vaccination. These range from low productivity of microbial fermentation to difficult downstream processing and lack of a platform process.The current study builds on the vast body of publications and expertise in the group at UCL to address a few of the bottlenecks in the manufacturability of plasmid DNA. Hypothesis: tightly formed supercoiled DNA can potentially overcome the challenges of low quantity and quality of large supercoiled plasmids produced at scale.The study focuses on the most recent research in our labs where we have been successful in demonstrating that we can create tight supercoiled DNA with reduced size. This has been achieved in 6.5 Kb plasmids and we will extend this to larger plasmids (beyond 20Kb) where more information can be packaged. We will then examine the challenges of production in the fermenter followed by recovery and initial separation steps.