双磁珠法质粒DNA提取技术及应用

Plasmid DNA extraction by double magnetic beads method and its application

摘要:

质粒是基因合成与测序领域中使用最为频繁的基因运载工具，然而传统的质粒DNA提取方法面临提取通量低、生产成本高等问题，无法满足日益增长的需求。本研究基于质粒提取原理，开发了双磁珠法(double-magnetic-bead method, DMBM)质粒提取技术，探究了磁珠投入量、质粒DNA片段大小、菌液投入量等因素对质粒提取的影响，并且对比了本技术与商业化质粒DNA提取试剂盒提取DNA质量、提取通量及提取成本。结果表明，双磁珠法质粒DNA提取技术可满足不同细胞密度、不同片段长度的质粒DNA提取。此外，该技术搭载96通道全自动核酸提取仪，提取的质粒DNA纯度更高、提取时间缩短80%、提取成本缩减57.1%，从而实现了质粒DNA提取的高通量、低成本，有效助力基因合成与测序。

Abstract:

Plasmids are the most commonly used gene carriers in the field of gene synthesis and sequencing. However, the main problems faced by traditional plasmid DNA extraction technology are low extraction throughput and high production cost, so they cannot meet the growing demand. In this study, a double-magnetic-bead method (DMBM) for plasmid extraction was developed based on the principle of plasmid extraction. The effects of the input of magnetic beads, the size of plasmid DNA fragments, and the volume of bacterial on plasmid DNA extraction were explored. In addition, the quality, throughput, and cost of plasmid DNA extraction were also compared between this technique and the commercial plasmid DNA extraction kits. The results showed that the DMBM can meet the needs of extracting plasmid DNA with different cell densities and fragment lengths. Moreover, the sensitivity and quality of plasmid extraction by the DMBM method were both superior to those of the centrifugal adsorption column method. In addition, this technique could be applied on a 96-channel automated nucleic acid extractor, resulting in higher purity of the extracted plasmid DNA, 80% reduction in extraction time, and 57.1% reduction in cost. It also reduces manual operations, achieving high-throughput and low-cost plasmid DNA extraction, thus may facilitate gene synthesis and sequencing.