Summary Molecular taxonomy is a field that studies the diversity of organisms based on molecular markers. This work is devoted to develop a methodology of molecular taxonomy of small organisms. The ribosomal RNA (rRNA) is used as a molecular marker since its nucleotide sequence includes stretches of various levels of conservation, which can be used as species, genus and taxa specific regions. The organisms live in complex communities. To discover the composition of these communities, a hybridization assay employing oligonucleotide microarrays is developed to indicate the presence of a certain rRNA, in a sample under investigation. An additional method based on the pyrosequencing process is proposed here. In this case the mixture of rRNA genes is directly sequenced and the proportion of individual sequences is then calculated from the obtained pyrogram. The work comprises two parts: theoretical bioinformatics and practical evaluation. The first part tackles the problem of DNA-RNA duplex stability prediction. As a result, an ad hoc stability function is proposed. An algorithm and a program are developed for the design of oligonucleotides employed in the microarray approach. The kinetics of DNA-RNA duplex dissociation is considered as well. In addition, the formalism of the pyrosequencing approach is elaborated theoretically. The experimental part deals with the issues of oligonucleotide microarray establishment, including fabrication, immobilization, hybridization and scanning. A real-time kinetic setup for observing the RNA-DNA duplex dissociation was developed. The theoretical findings and quality of the oligonucleotide design are practically evaluated. The theory is found to be in a good accordance with experiment. The pyrosequencing approach is tested as well and is demonstrated to have enough power to discover the composition of a complex mixture of rRNA genes
