Short DNA fragments containing single, uniquely positioned nucleosome cores have been extensively employed as simple model experimental systems for analysis of many intranuclear processes, including binding of proteins to nucleosomes, transcription, DNA repair and ATP-dependent chromatin remodeling. In many cases such simple model templates faithfully recapitulate numerous important aspects of these processes. Here we describe several recently developed procedures for obtaining and analysis of mononucleosomes that are uniquely positioned on 150-600 bp DNA fragments.