Real-time polymerase chain reaction assay for rapid and sensitive detection of anthrax spores in spiked soil and talcum powder
Language English Country United States Media print-electronic
Document type Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- Bacillus anthracis genetics isolation & purification MeSH
- Bacteriological Techniques methods MeSH
- Time Factors MeSH
- DNA, Bacterial genetics isolation & purification MeSH
- Real-Time Polymerase Chain Reaction methods MeSH
- Talc * MeSH
- Soil Microbiology * MeSH
- Sensitivity and Specificity MeSH
- Spores genetics isolation & purification MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- DNA, Bacterial MeSH
- Talc * MeSH
Real-time polymerase chain reaction (real-time PCR) is a laboratory technique based on PCR. This technique is able to detect sequence-specific PCR products as they accumulate in "real time" during the PCR amplification, and also to quantify the number of substrates present in the initial PCR mixture before amplification begins. In the present study, real-time PCR assay was employed for rapid and real-time detection of Bacillus anthracis spores spiked in 0.1 g of soil and talcum powder ranging from 5 to 10(7) spores. DNA was isolated from spiked soil and talcum powder, using PBS containing 1 % Triton-X-100, followed by heat treatment. The isolated DNA was used as template for real-time PCR and PCR. Real-time PCR amplification was obtained in 60 min under the annealing condition at 60°C by employing primers targeting the pag gene of B. anthracis. In the present study, the detection limit of real-time PCR assay in soil was 10(3) spores and 10(2) spores in talcum powder, respectively, whereas PCR could detect 10(4) spores in soil and 10(3) spores in talcum powder, respectively.
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