Department of Molecular Biosciences, College of Natural Sciences, The University of Texas at Austin, Austin, Texas, United States of America.
Center for Systems and Synthetic Biology, The University of Texas at Austin, Austin, Texas, United States of America.
PLoS One. 2021 Jun 1;16(6):e0252507. doi: 10.1371/journal.pone.0252507. eCollection 2021.
We recently developed 'cellular' reagents-lyophilized bacteria overexpressing proteins of interest-that can replace commercial pure enzymes in typical diagnostic and molecular biology reactions. To make cellular reagent technology widely accessible and amenable to local production with minimal instrumentation, we now report a significantly simplified method for preparing cellular reagents that requires only a common bacterial incubator to grow and subsequently dry enzyme-expressing bacteria at 37°C with the aid of inexpensive chemical desiccants. We demonstrate application of such dried cellular reagents in common molecular and synthetic biology processes, such as PCR, qPCR, reverse transcription, isothermal amplification, and Golden Gate DNA assembly, in building easy-to-use testing kits, and in rapid reagent production for meeting extraordinary diagnostic demands such as those being faced in the ongoing SARS-CoV-2 pandemic. Furthermore, we demonstrate feasibility of local production by successfully implementing this minimized procedure and preparing cellular reagents in several countries, including the United Kingdom, Cameroon, and Ghana. Our results demonstrate possibilities for readily scalable local and distributed reagent production, and further instantiate the opportunities available via synthetic biology in general.
我们最近开发了“细胞”试剂——冻干过表达目的蛋白的细菌,可替代典型诊断和分子生物学反应中的商业纯酶。为了使细胞试剂技术广泛普及,并可通过最少的仪器在当地生产,我们现在报告一种显著简化的细胞试剂制备方法,该方法仅需一个常见的细菌培养箱,在 37°C 下用廉价的化学干燥剂帮助生长并随后干燥表达酶的细菌。我们展示了此类干燥细胞试剂在常见分子和合成生物学过程中的应用,如 PCR、qPCR、逆转录、等温扩增和 Golden Gate DNA 组装,用于构建易于使用的检测试剂盒,以及用于快速生产满足特殊诊断需求的试剂,例如当前正在应对的 SARS-CoV-2 大流行。此外,我们通过在包括英国、喀麦隆和加纳在内的几个国家成功实施这一简化程序并制备细胞试剂,证明了本地生产的可行性。我们的结果表明,本地和分布式试剂生产具有易于扩展的可能性,并进一步体现了一般合成生物学提供的机会。