Site directed in-vitro assembly of nucleosomes

An in-vitro system is described allowing for the assembly of nucleosomes on preselected sites of a cloned tRNA gene. The system consists of a soluble nucleoprotein fraction, a histone source, and circular DNA containing a single stranded stretch (sssDNA). Nucleosomes assemble on the sssDNA, if the three components are incubated as a highly concentrated solution in the presence of the four deoxyribonucleotidetriphosphates and of ATP. The single stranded stretch is rendered double stranded during incubation. The middle axis of one assembled nucleosome always coincides roughly with the midpoint of the original single stranded DNA stretch.