Journal
JOURNAL OF CHROMATOGRAPHY A
Volume 1548, Issue -, Pages 100-103Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.chroma.2018.03.021
Keywords
DNA; lsotachophoresis; Nucleic acids; Sample preparation
Funding
- Grant Agency of the Czech Republic [P20612G014, MTB 8F17003]
- Roche Sequencing Solutions (Pleasanton, USA)
- Institute of Analytical Chemistry, Czech Academy of Sciences, v.v.i. (Brno, Czech Republic) [RVO 68081715]
- Ministry of Education, Youth and Sports of the Czech Republic under the National Sustainability Programme II [LQ1601]
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Sample preparation plays an important role in the DNA analysis workflow. Real samples often include a complex matrix, such as blood and other bodily fluids, or exogenous impurities, e.g., from the scene of crime. Most of the common nucleic acids isolation techniques are based on extractions: however, isotachophoretic focusing has recently attracted some interest for its simplicity and potential for very high enrichment factors and ease of automation, Here, we report on the use of a commercial isotachophoretic instrument for optimization of DNA focusing and preparative fraction collection. In order to achieve a high recovery and enrichment, experimental factors including electric current, sample amount and matrix were investigated experimentally as well as by computer simulation. The sample of a DNA ladder was injected in 30 volume and after ITP focusing the DNA zone was recovered using an on-column micropreparative collection valve. The DNA content in the collected sample was verified by fluorescence spectrometry and chip capillary electrophoresis with fluorescence detection. (C) 2018 Elsevier B.V. All rights reserved.
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