Comparative analysis of the efficiency of methods for DNA extraction from animal tissues

Abstract

For the first time, a comparative analysis of the effectiveness of DNA extraction methods used in expert forensic practice from various animal tissues – bone antlers and hooves of reindeer Rangifer tarandus (Linnaeus, 1758); skeletal muscle, cartilage tissue and mucus of rainbow trout Oncorhynchus mykkis (Walbaum, 1792); skeletal muscles and ribs of the domestic pig Sus scrofa domesticus (Erxleben, 1777). DNA extraction was carried out according to the manufacturers' protocols using six commercial reagent kits, domestically produced, based on the alcohol precipitation method: "GM Tissue" (Raissol), “Amex” (Raissol); sorption onto magnetic particles: “GM Tissue M” (“Raissol”), “M-Sorb-bone” (“Synthol”); sorption on silica gel: “D-Tissues” (“Biolabmix”), “D-blood” (“Biolabmix”). The quantitative and qualitative characteristics of the obtained DNA were assessed by measuring the concentration of double-stranded DNA on a Qubit 2.0 fluorometer (Life Technologies, USA) using the Qubit DNA HS Assay Kit. The highest concentration of double-stranded DNA of all types of animal tissue was obtained by sorption onto magnetic particles. Using this method, the most stable and reproducible DNA concentrations were obtained (average values – 474.00 μg/ml, 268.00 μg/ml, 83.20 μg/ml and 69.70 μg/ml, respectively). Using the method of DNA extraction based on precipitation with alcohol, lower concentrations of DNA were obtained from all types of animal tissues in comparison with the method of sorption onto magnetic particles. However, the use of this method has shown a fairly high stability and reproducibility of the obtained DNA concentrations (average values – 120.00 µg/ml, 110.00 µg/ml and 99.90 µg/ml, respectively), from all types of biological tissues, in all series (repetitions) of the extraction step. The use of a DNA extraction method based on sorption on silica gel showed, on average, lower concentrations of target DNA from all types of animal tissues, in comparison with the methods of sorption on magnetic particles and precipitation with alcohol. This method was characterized by the lowest stability and reproducibility of the obtained DNA concentrations (average values – 74.10 μg/ml and 41.60 μg/ml, respectively). Thus, the team of authors assessed the effectiveness of methodological approaches to DNA extraction in relation to various types of animal tissue, as well as assessed the stability and reproducibility of the results obtained. The effectiveness of these methodological approaches has not been previously assessed.