Påvisning av biologiske stridsmidler ved bruk av real-time PCR

Genetic methods are nowadays considered indispensable for identification of microorganisms. Such methods are used in the field of food safety, clinical diagnostics of humans and animals and for identification of biological warfare agents. In PCR, specific DNA-sequences are amplified and these PCR products are detected by gel electrophoresis. In real-time PCR amplified DNA is detected during the PCR by using the fluorescent dye SYBR Green I or fluorescent labelled DNA probes. Real-time PCR has considerable advantages over classical PCR. It is fast (20-40 minutes), possibility of quantification and no additional detection method such as gel electrophoresis is needed, which means decreased risk of contamination between different samples. In this report we describe real- time PCR methods for identification of several biological warfare agents such as Bacillus anthracis, Yersinia pestis, Vibrio cholerae, Coxiella burnetii, Francisella tularensis and Brucella melitensis. A method for analyisis of Vaccinia virus is also included. These samples are provided by Dr Bruce Harper, Dugway Proving Ground, Life Science Division, USA. In addition we shortly describe a method for lysis of Gram-positive bacteria exampled by Bacillus cereus.