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We investigated the effect of tris(hydroxymethyl)aminomethane hydrochloride (Tris-HCl) buffer (pH 7.0) as a bulk solution on the adsorption of DNA by gibbsite, goethite, montmorillonite, kaolinite, synthetic and natural allophanes, two humic acids and two andosols. The natural allophane, gibbsite, kaolinite and an andosol adsorbed significantly more DNA in a 0.1 M Tris-HCl buffer than in a 0.1 M NaCl solution (t-test, P<0.005). In contrast, montmorillonite adsorbed significantly less DNA in the Tris-HCl than NaCl solution (P<0.05). Care should be taken when using TrisHCl in studies on the adsorption of extracellular DNA molecules by soil particles.Key words: DNA adsorption, Tris-HCl buffer, allophane, soil, montmorilloniteNucleic acid adsorption at soil-solution interfaces is important in understanding many subjects such as the biosafety of genetically modified organisms (17), protection of DNA against nucleases in soil (13, 22), genetic transformation of competent bacteria in situ (5,6,11,22) and extraction of DNA from soil for the study of microbial communities (e.g. 34).The adsorption of DNA molecules by phyllosilicate minerals is affected by several factors including DNA size (6), mineral types (2), the pH of buffer solutions (7,8,11), the ionic strength of solutions (12), the presence of other cellular components (21, 22) and electrolytes (7). The adsorption of DNA by phyllosilicate minerals can differ from that by variable-charge soils such as andosol. There have been several studies on the retention of DNA in natural soils (18,19,26,28,29).Tris(hydroxymethyl)aminomethane (Tris) solutions are used widely as buffers due to their stabilizing effects on biomolecules (16). There have been several studies about the adsorption of DNA on soil particles using Tris-HCl buffers as background solutions (e.g. 2, 3, 4). Recently, Wei et al. (36) reported that a Tris-HCl buffer (pH 7.4) increased protein adsorption on germanium (Ge) surface. To grasp how much Tris-HCl affects on the adsorption of DNA by soils, we investigated the effect of a Tris-HCl buffer (pH 7.0) as a bulk solution on DNA adsorption by a variety of materials (gibbsite, goethite, montmorillonite, kaolinite, synthetic and natural allophanes, two humic acids and two andosols).The mineral samples used in this study were: Gibbsite, provided by Showa Denko (Tokyo, Japan); goethite, synthesized by the method of Atkinson et al.(1); montmorillonite K10, provided by Sigma-Aldrich (St Louis, USA); kaolinite, collected from Iriki, Kagoshima, Japan; a synthetic allophane produced by the method of Ohashi et al. (20) and a natural allophane supplied by Shinagawa Chemicals (Tokyo, Japan). The properties of these solids have been reported recently (14,24,25). Humic acid A and B were purified from reagents provided by Wako Pure Chemical Industries (Osaka, Japan) and Sigma-Aldrich. The purification process was based on the IHSS method (33). The properties of humic acids were described previously (10,15,27). Two surface humic andosols were also used; Chiran soil (CR ...
We investigated the effect of tris(hydroxymethyl)aminomethane hydrochloride (Tris-HCl) buffer (pH 7.0) as a bulk solution on the adsorption of DNA by gibbsite, goethite, montmorillonite, kaolinite, synthetic and natural allophanes, two humic acids and two andosols. The natural allophane, gibbsite, kaolinite and an andosol adsorbed significantly more DNA in a 0.1 M Tris-HCl buffer than in a 0.1 M NaCl solution (t-test, P<0.005). In contrast, montmorillonite adsorbed significantly less DNA in the Tris-HCl than NaCl solution (P<0.05). Care should be taken when using TrisHCl in studies on the adsorption of extracellular DNA molecules by soil particles.Key words: DNA adsorption, Tris-HCl buffer, allophane, soil, montmorilloniteNucleic acid adsorption at soil-solution interfaces is important in understanding many subjects such as the biosafety of genetically modified organisms (17), protection of DNA against nucleases in soil (13, 22), genetic transformation of competent bacteria in situ (5,6,11,22) and extraction of DNA from soil for the study of microbial communities (e.g. 34).The adsorption of DNA molecules by phyllosilicate minerals is affected by several factors including DNA size (6), mineral types (2), the pH of buffer solutions (7,8,11), the ionic strength of solutions (12), the presence of other cellular components (21, 22) and electrolytes (7). The adsorption of DNA by phyllosilicate minerals can differ from that by variable-charge soils such as andosol. There have been several studies on the retention of DNA in natural soils (18,19,26,28,29).Tris(hydroxymethyl)aminomethane (Tris) solutions are used widely as buffers due to their stabilizing effects on biomolecules (16). There have been several studies about the adsorption of DNA on soil particles using Tris-HCl buffers as background solutions (e.g. 2, 3, 4). Recently, Wei et al. (36) reported that a Tris-HCl buffer (pH 7.4) increased protein adsorption on germanium (Ge) surface. To grasp how much Tris-HCl affects on the adsorption of DNA by soils, we investigated the effect of a Tris-HCl buffer (pH 7.0) as a bulk solution on DNA adsorption by a variety of materials (gibbsite, goethite, montmorillonite, kaolinite, synthetic and natural allophanes, two humic acids and two andosols).The mineral samples used in this study were: Gibbsite, provided by Showa Denko (Tokyo, Japan); goethite, synthesized by the method of Atkinson et al.(1); montmorillonite K10, provided by Sigma-Aldrich (St Louis, USA); kaolinite, collected from Iriki, Kagoshima, Japan; a synthetic allophane produced by the method of Ohashi et al. (20) and a natural allophane supplied by Shinagawa Chemicals (Tokyo, Japan). The properties of these solids have been reported recently (14,24,25). Humic acid A and B were purified from reagents provided by Wako Pure Chemical Industries (Osaka, Japan) and Sigma-Aldrich. The purification process was based on the IHSS method (33). The properties of humic acids were described previously (10,15,27). Two surface humic andosols were also used; Chiran soil (CR ...
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Environmental DNA (eDNA) once shed can exist in numerous states with varying behaviors including degradation rates and transport potential. In this study, we consider three states of eDNA: (1) a membrane-bound state referring to DNA enveloped in a cellular or organellar membrane, (2) a dissolved state defined as the extracellular DNA molecule in the environment without any interaction with other particles, and(3) an adsorbed state defined as extracellular DNA adsorbed to a particle surface in the environment. Capturing, isolating, and analyzing a target state of eDNA provides utility for better interpretation of eDNA degradation rates and transport potential.While methods for separating different states of DNA have been developed, they remain poorly evaluated due to the lack of state-controlled experimentation. We evaluated the methods for separating states of eDNA from a single sample by spiking DNA from three different species to represent the three states of eDNA as state-specific controls. We used chicken DNA to represent the dissolved state, cultured mouse cells for the membrane-bound state, and salmon DNA adsorbed to clay particles as the adsorbed state. We performed the separation in three water matrices, two environmental and one synthetic, spiked with the three eDNA states. The membrane-bound state was the only state that was isolated with minimal contamination from nontarget states. The membrane-bound state also had the highest recovery (54.11 ± 19.24%), followed by the adsorbed state (5.08 ± 2.28%), and the dissolved state had the lowest total recovery (2.21 ± 2.36%). This study highlights the potential to sort the states of eDNA from a single sample and independently analyze them for more informed biodiversity assessments. However, further method development is needed to improve recovery and reduce cross-contamination.
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