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利用多种化学化合物和纳米材料优化各种细菌物种的理化转化方法。

Optimization of chemico-physical transformation methods for various bacterial species using diverse chemical compounds and nanomaterials.

机构信息

School of Integrative Engineering, Chung-Ang University, Seoul, 06974, Republic of Korea.

School of Integrative Engineering, Chung-Ang University, Seoul, 06974, Republic of Korea.

出版信息

J Biotechnol. 2018 Dec 20;288:55-60. doi: 10.1016/j.jbiotec.2018.11.003. Epub 2018 Nov 5.

Abstract

Bacterial transformation is a fundamental technology to deliver engineered plasmids into bacterial cells, which is essential in industrial protein production, chemical production, etc. Previously, we developed a simple chemico-physical transformation method that can be applied to various bacterial species. Here, to accelerate the advance of bacteria biotechnology we optimize our method by combinatorially evaluating chemical compounds (rubidium chloride, lithium acetate, cesium chloride, dimethyl sulfoxide, and magnesium chloride) for increasing membrane permeability and nanomaterials (sepiolite, gold(III) chloride, multiwalled carbon nanotube, and chitosan) for piercing the membranes. The best transformation efficiencies were achieved as follows; 2.84 × 10 CFU/μg DNA in Lactococcus lactis subsp. lactics (0.1 M CsCl and gold(III) chloride), 3.60 × 10 CFU/μg DNA in Enterococcus faecalis (1 M Li-acetate and MWCNT), 2.41 × 10 CFU/μg DNA in Bacillus sp. (0.01 M RbCl and sepiolite), 3.49 × 10 CFU/μg DNA (0.1 M RbCl and gold(III) chloride) in Ralstonia eutropha (also known as Cupriavidus necator) and 8.78 × 10 CFU/μg DNA (1 M RbCl and chitosan) in Methylomonas sp. DH-1. The efficiencies are up to 100-fold higher than those without optimization. Accordingly, our fast and simple chemico-physical transformation with chemical-nanomaterial optimization allows for the efficient DNA entry into various bacterial cells with high efficiency.

摘要

细菌转化是将工程质粒导入细菌细胞的基础技术,在工业蛋白质生产、化学生产等方面至关重要。此前,我们开发了一种简单的理化转化方法,可应用于各种细菌。在这里,为了加速细菌生物技术的进步,我们通过组合评估化学化合物(氯化铷、醋酸锂、氯化铯、二甲基亚砜和氯化镁)来提高细胞膜通透性和纳米材料(海泡石、三氯化金、多壁碳纳米管和壳聚糖)来刺穿细胞膜,对我们的方法进行了优化。最佳转化效率如下:乳酸乳球菌亚种(0.1 M CsCl 和三氯化金)为 2.84×10 CFU/μg DNA,粪肠球菌(1 M Li-acetate 和 MWCNT)为 3.60×10 CFU/μg DNA,芽孢杆菌(0.01 M RbCl 和海泡石)为 2.41×10 CFU/μg DNA,罗尔斯通氏菌(也称为醋杆菌属)为 3.49×10 CFU/μg DNA(0.1 M RbCl 和三氯化金),甲基单胞菌 DH-1 为 8.78×10 CFU/μg DNA(1 M RbCl 和壳聚糖)。效率比没有优化时提高了 100 倍。因此,我们的快速、简单的化学物理转化与化学纳米材料优化相结合,允许高效的 DNA 进入各种细菌细胞,效率很高。

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