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Efficiency of DNA Isolation Methods Based on Silica Columns and Magnetic Separation Tested for the Detection of Mycobacterium avium Subsp. Paratuberculosis in Milk and Faeces.
Husakova, Marketa; Kralik, Petr; Babak, Vladimir; Slana, Iva.
Afiliação
  • Husakova M; Veterinary Research Institute, Hudcova 70, 621 00 Brno, Czech Republic.
  • Kralik P; Department of Experimental Biology, Faculty of Science, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.
  • Babak V; Veterinary Research Institute, Hudcova 70, 621 00 Brno, Czech Republic.
  • Slana I; Department of Hygiene and Technology of Food of Animal Origin and of Gastronomy, Faculty of Veterinary Hygiene and Ecology, University of Veterinary and Pharmaceutical Sciences, 612 42 Brno, Czech Republic.
Materials (Basel) ; 13(22)2020 Nov 12.
Article em En | MEDLINE | ID: mdl-33198402
Timely and reliable detection of animals shedding Mycobacterium avium subsp. paratuberculosis (MAP) should help to effectively identify infected animals and limit infection transmission at early stages to ensure effective control of paratuberculosis. The aim of the study was to compare DNA extraction methods and evaluate isolation efficiency using milk and faecal samples artificially contaminated by MAP with a focus on modern instrumental automatic DNA isolation procedures based on magnetic separation. In parallel, an automatic and manual version of magnetic separation and two methods of faecal samples preparation were compared. Commercially available DNA isolation kits were evaluated, and the selected kits were used in a trial of automatic magnetic beads-based isolation and compared with the manual version of each kit. Detection of the single copy element F57 was performed by qPCR to quantify MAP and determine the isolation efficiency. The evaluated kits showed significant differences in DNA isolation efficiencies. The best results were observed with the silica column Blood and Tissue kit for milk and Zymo Research for faeces. The highest isolation efficiency for magnetic separation was achieved with MagMAX for both matrices. The magnetic separation and silica column isolation methods used in this study represent frequently used methods in mycobacterial diagnostics.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Tipo de estudo: Diagnostic_studies / Guideline Idioma: En Revista: Materials (Basel) Ano de publicação: 2020 Tipo de documento: Article País de afiliação: República Tcheca País de publicação: Suíça

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Tipo de estudo: Diagnostic_studies / Guideline Idioma: En Revista: Materials (Basel) Ano de publicação: 2020 Tipo de documento: Article País de afiliação: República Tcheca País de publicação: Suíça