Vándor Andrea szerk.: Janus Pannonius Múzeum Évkönyve 50-52/2 (2005-2007) (Pécs, 2008)

Albert Zink: Molecular results of the analysis of the sample Pa

Albert Zink Molecular results of the analysis of the sample Pa 6 EURAC - Institute for Mummies and the Iceman Bolzano, Italy A small bone sample of the case Pa 6 was investigated together with other Hungarian samples within the scope of bilateral collaboration of our two institutes supported by the DAAD (German Academic Exchange Service). The sample was superficially decontamined and subsequently subjected to DNA extraction, amplification and analyzing procedures following wellestablished standard protocols in our labs. In a first step, a 202bp fragment of the human ß-actin gene was amplified to test whether DNA was successfully extracted from the specimen. This amplification provided a clear signal at the expected size in the gel electrophoresis. A molecular sex determination by amplification of the amelogenin gene provided no amplicication product. This analysis will be repeated. In the next step, the sample was tested for the presence of Mycobacterium tuberculosis complex DNA. For this a 123bp segment of the IS6110 was amplified, which is highly specific for the Mycobacterium tuberculosis complex. The PCR amplification of the IS6110 was repeated several times. In parallel, this analytic procedure was completed by a two-round nested PCR, where the amplification product of the IS6110 was subjected to a second PCR amplifying a 92bp fragment lying within the 123bp IS6110 product. In both amplifications a single band of the expected size was detected. For further testing the specificity of the Mycobacterium tuberculosis specific amplification, the 123bp PCR product was digested by the restriction enzyme Hae III, resulting in a 94bp and 29bp fragment. The restriction enzyme digestion confirmed the positive results of the PCR amplification of sample Pa 6. In an additional analytic step, the Mycobacterium tuberculosis DNA of specimen Pa 6 was further characterized by spoligotyping. This technique allows the differentiation of the members of the Mycobacterium tuberculosis complex (M. tuberculosis, M. bovis, M. africanum, M. microti and M. canetti) and provides information about their affilitation to a certain mycobacterial strain. This is from particular interest for the evaluation of evolutionary processes and the distribution of tuberculosis in populations over different times and geographical regions. The spoligotyping resulted in an incomplete pattern, which permitted the determination of the species, but no detailed classification to a certain strain. The spoligotyping method provided, in considetarion of the incomplete hybridization, atypical Mycobacterium tuberculosis signature. In the near future, the analysis will be repeated in oder to get a more complete pattern. A Janus Pannonius Múzeum Évkönyve 1 151