The Use of PCR to Detect a Pathogen in Compost |
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| Authors: | Stacy L Pfaller Stephen J Vesper Hector Moreno |
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| Institution: | 1. Department of Civil and Environmental Engineering, University of Cincinnati, Cincinnati, Ohio;2. U. S. Environmental Protection Agency, Center Hill Laboratory, Cincinnati, Ohio |
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| Abstract: | Testing compost for pathogens can be very difficult, time consuming and expensive. A method to purify DNA from compost for amplification by polymerase chain reaction (PCR) was sought. The published techniques used to purify DNA from soils or sediments did not work with compost samples in our laboratory. We have devised an extraction protocol specifically for compost samples. In a series of Escherichia coli-spiked compost samples, ranging from 108 to 103 organisms per gram of compost, the removal of substances that inhibit PCR amplification of DNA was accomplished by extracting the organisms from the compost and treating the extract with base resin. After cell lysis, the collected DNA was further purified on a 1 percent low melt agarose gel. A 260 base-pair region of the lac Z gene, unique to E. coli, was successfully amplified by PCR in the DNA extracted from the gel. The technique was sensitive to the level of 104 E. coli cells per gram of compost. |
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