National Institute of Standards and Technology, Gaithersburg, MD, United States of America.
Department of Chemistry and Biochemistry, University of Maryland, College Park, MD, United States of America.
PLoS One. 2023 Oct 17;18(10):e0292401. doi: 10.1371/journal.pone.0292401. eCollection 2023.
Although many protocols have been previously developed for genomic DNA (gDNA) extraction from S. cerevisiae, to take advantage of recent advances in laboratory automation and DNA-barcode sequencing, there is a need for automated methods that can provide high-quality gDNA at high efficiency. Here, we describe and demonstrate a fully automated protocol that includes five basic steps: cell wall and RNA digestion, cell lysis, DNA binding to magnetic beads, washing with ethanol, and elution. Our protocol avoids the use of hazardous reagents (e.g., phenol, chloroform), glass beads for mechanical cell disruption, or incubation of samples at 100°C (i.e., boiling). We show that our protocol can extract gDNA with high efficiency both from cells grown in liquid culture and from colonies grown on agar plates. We also show results from gel electrophoresis that demonstrate that the resulting gDNA is of high quality.
虽然之前已经开发了许多用于从酿酒酵母中提取基因组 DNA（gDNA）的方案，但为了利用实验室自动化和 DNA 条码测序的最新进展，需要开发能够高效提供高质量 gDNA 的自动化方法。在这里，我们描述并验证了一种完全自动化的方案，该方案包括五个基本步骤：细胞壁和 RNA 消化、细胞裂解、DNA 与磁珠结合、乙醇洗涤和洗脱。我们的方案避免了使用危险试剂（如苯酚、氯仿）、用于机械细胞破碎的玻璃珠或在 100°C 下孵育样品（即煮沸）。我们表明，我们的方案可以从液体培养中生长的细胞和琼脂平板上生长的菌落中高效提取 gDNA。我们还展示了凝胶电泳的结果，证明了所得 gDNA 的高质量。
