DIRECT DNA EXTRACTION FOR PCR-MEDIATED ASSAYS OF SOIL ORGANISMS

By using the rDNA of a plant wilt pathogen (Verticillium dahliae) as t he target sequence, a direct method for the extraction of DNA from soi l samples which can be used for PCR-mediated diagnostics without a nee d for further DNA purification has been developed. The soil organisms are disrupted by grinding in liquid nitrogen with the natural abrasive s in soil, and losses due to degradation and adsorption are largely el iminated by the addition of skim milk powder. The DNA from disrupted c ells is extracted with sodium dodecyl sulfate-phenol and collected by ethanol precipitation. After suitable dilution, this DNA extract can b e assayed directly by PCR amplification technologies. The method is ra pid, cost efficient, and when combined with suitable internal controls can be applied to the detection and quantification of specific soil o rganisms or pathogens on a large-scale basis.