Principles and practice of nucleosome positioning in vitro

Understanding the function of chromatin has become an exciting field that interests researchers from a wide variety of disciplines. Chromatin is composed of fundamental nucleosome building blocks, and the ability to assemble defined, positioned mononucleosomes is central to experimental investigation of chromatin structure, stability and dynamics. However, the molecular principles, historical background and biochemistry of experimental techniques underlying positioned nucleosomes are sometimes unclear. This review provides an introduction from the perspective of the core histone octamer as a surface for histone-DNA interactions that directs a unique conformation for wrapped DNA. The origins and properties of the most widely used nucleosome positioning DNA sequences including the Widom 601, 5S rRNA and MMTV LTR as well as other less well-known examples are described. An overview of the main experimental methods for preparing nucleosomes and mapping their positions is also provided. These should be suitable for researchers entering the field of chromatin and enable an appreciation of the principles and practical aspects of experimental investigations using positioned nucleosomes in vitro.