Fortin Nathalie, Beaumier Danielle, Lee Kenneth, Greer Charles W
Biotechnology Research Institute, National Research Council of Canada, 6100 Royalmount Ave., Montreal, QC, Canada H4P 2R2.
J Microbiol Methods. 2004 Feb;56(2):181-91. doi: 10.1016/j.mimet.2003.10.006.
Treatment of soil with surfactants and chelating agents is used in bioremediation studies to desorb and solubilize contaminants to increase their bioavalability to microorganisms. In the same way that pollutants are made more bioavailable to microorganisms, the procedure can be used to remove potential interfering materials from soil prior to cell lysis and extraction of DNA from indigenous microorganisms. The effect of soil washing was evaluated by extracting DNA from sediments of an intertidal freshwater wetland contaminated with hydrocarbons and from highly contaminated marine sediments from Sydney Harbour, Nova Scotia, Canada. Sediment samples had total organic carbon (TOC) contents that varied between 0.2% and 13%. The chemical lysis technique was also examined by comparison of an ammonium acetate precipitation of proteins and humic acids with a hexadecyltrimethylammonium bromide (CTAB) incubation and phenol:chloroform extraction. In this study, the incorporation of soil washing steps facilitated the desorption of contaminants from sediment surfaces and improved the recovery of DNA of amplifiable quality from both freshwater and marine sediments. CTAB contributed only slightly to the recovery of DNA of higher quality in the most contaminated sample from Sydney Harbour and was concomitant with a decrease in DNA yield in both sediment types. The incorporation of a soil washing step prior to the extraction of DNA from polluted environments may be important to solubilize and remove contaminants when high-quality DNA is required for subsequent analyses.
在生物修复研究中,使用表面活性剂和螯合剂处理土壤,以解吸和溶解污染物,提高其对微生物的生物可利用性。与使污染物对微生物更具生物可利用性的方式相同,该程序可用于在细胞裂解和从本地微生物中提取DNA之前,从土壤中去除潜在的干扰物质。通过从受碳氢化合物污染的潮间带淡水湿地沉积物以及加拿大新斯科舍省悉尼港高度污染的海洋沉积物中提取DNA,评估了土壤洗涤的效果。沉积物样品的总有机碳(TOC)含量在0.2%至13%之间变化。还通过比较用十六烷基三甲基溴化铵(CTAB)孵育和苯酚:氯仿提取法沉淀蛋白质和腐殖酸的醋酸铵沉淀法,研究了化学裂解技术。在本研究中,加入土壤洗涤步骤有助于从沉积物表面解吸污染物,并提高了从淡水和海洋沉积物中回收可扩增质量DNA的效率。在悉尼港污染最严重的样品中,CTAB对高质量DNA回收的贡献仅略大,并且两种沉积物类型中的DNA产量均随之下降。当后续分析需要高质量DNA时,在从污染环境中提取DNA之前加入土壤洗涤步骤对于溶解和去除污染物可能很重要。