Institute of Medical Biotechnology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
Institute of Medical Biotechnology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
Biosens Bioelectron. 2019 Apr 15;131:95-103. doi: 10.1016/j.bios.2019.02.024. Epub 2019 Feb 19.
Electroporation has been a widely established method for delivering DNA and other material into cells in vitro. Conventional electroporation infrastructure is typically immobile, non-customizable, non-transparent regarding the characteristics of output pulses, and expensive. Here, we describe a portable electroporator for DNA delivery into bacterial cells that can quickly be reconstructed using 3D desktop printing and off-the-shelf components. The device is light weight (700 g), small (70 × 180 × 210 mm) and extremely low-cost (<EUR 130). We provide the electrical circuitry and a detailed parts list for rebuilding the device. We characterize the properties of generated pulses and apply the system for gene delivery into bacterial Dh5α cells. We analyze the transformation efficiency based on the optical density of cell suspensions at 595 nm and on quantitative analysis of images obtained from bacterial cell-grown agar plates using colony forming units as well as confluence as indicators. We demonstrate time-dependency of the transformation efficiency using single pulses of 500 V between 1 and 1000 ms duration and we show that commercially available electroporation cuvettes of 1 mm gap size reveal higher transformation rates compared to cuvettes with 2 mm gap. We benchmark the transformation efficiency obtained using our platform with data from a heat shock-based transformation protocol and with data from a commercially available electroporator and show that our system reveals comparable results as the other techniques in the applied configurations. While this work focuses on genetic manipulation of bacterial cells, the device may also be applicable for delivery of genetic material small molecule or nanomaterials into other cell types, including mammalian cells.
电穿孔已被广泛应用于将 DNA 和其他物质递送入体外细胞。传统的电穿孔基础设施通常是固定的、不可定制的、对输出脉冲特性不透明的,而且价格昂贵。在这里,我们描述了一种用于将 DNA 递送入细菌细胞的便携式电穿孔仪,它可以使用 3D 桌面打印和现成的组件快速重建。该设备重量轻(700 克)、体积小(70×180×210 毫米)、成本极低(<EUR 130)。我们提供了用于重建设备的电子电路和详细的零件清单。我们对生成的脉冲特性进行了表征,并将该系统应用于基因递送入细菌 Dh5α 细胞。我们基于细胞悬浮液在 595nm 处的光密度以及通过使用菌落形成单位和汇合度作为指标从细菌细胞生长琼脂平板获得的图像的定量分析来分析转化效率。我们通过使用 500V 的单次脉冲(持续时间为 1 至 1000ms)来研究转化效率的时间依赖性,并且表明 1mm 间隙尺寸的商用电穿孔管比 2mm 间隙尺寸的管具有更高的转化效率。我们使用我们的平台获得的转化效率与基于热休克的转化方案的数据以及与商用电穿孔仪的数据进行基准比较,并表明我们的系统在应用配置中与其他技术具有可比的结果。虽然这项工作侧重于细菌细胞的遗传操作,但该设备也可适用于将遗传物质、小分子或纳米材料递送入其他细胞类型,包括哺乳动物细胞。