Hydrophobic ionic liquids for quantitative bacterial cell lysis with subsequent DNA quantification

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• 作者：Sabine Fuchs-Telka ; Susanne Fister…
• 关键词：Ionic liquids ; DNA extraction ; Quantitative PCR ; Salmonella Typhimurium ; Escherichia coli ; Solvent extraction ; Two ; phase system
• 刊名：Analytical and Bioanalytical Chemistry
• 出版年：2017
• 出版时间：February 2017
• 年：2017
• 卷：409
• 期：6
• 页码：1503-1511
• 全文大小：1429KB
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Analytical Chemistry; Biochemistry, general; Laboratory Medicine; Characterization and Evaluation of Materials; Food Science; Monitoring/Environmental Analysis;
• 出版者：Springer Berlin Heidelberg
• ISSN：1618-2650
• 卷排序：409

文摘

DNA is one of the most frequently analyzed molecules in the life sciences. In this article we describe a simple and fast protocol for quantitative DNA isolation from bacteria based on hydrophobic ionic liquid supported cell lysis at elevated temperatures (120–150 °C) for subsequent PCR-based analysis. From a set of five hydrophobic ionic liquids, 1-butyl-1-methylpyrrolidinium bis(trifluoromethylsulfonyl)imide was identified as the most suitable for quantitative cell lysis and DNA extraction because of limited quantitative PCR inhibition by the aqueous eluate as well as no detectable DNA uptake. The newly developed method was able to efficiently lyse Gram-negative bacterial cells, whereas Gram-positive cells were protected by their thick cell wall. The performance of the final protocol resulted in quantitative DNA extraction efficiencies for Gram-negative bacteria similar to those obtained with a commercial kit, whereas the number of handling steps, and especially the time required, was dramatically reduced.