The awareness that global biological diversity is affected by numerous, mostly human-made threats has made biodiversity assessment an important scientific issue for decades. Biodiversity includes different levels of complexity, such as community diversity, habitat diversity, genetic diversity, and species diversity. The application of molecular methods to answer ecological questions permits issues of biodiversity to be addressed at all levels. Microorganisms dominate global biological diversity in terms of their species numbers. However, their small size and limited morphological features make it challenging to obtain a comprehensive view of their biodiversity. The application of ribosomal RNA (rRNA) probes contributes significantly to the assessment of biodiversity at the molecular level. DNA microarrays offer a great potential to facilitate the application of molecular probes and other DNA analytical methods to answer ecological and biodiversity questions. We provide an introduction into the application of rRNA probes and DNA microarrays for the assessment of microbial biodiversity, as well as protocols for the implementation of DNA microarrays.