Development of methodology for studying chromatin fibers in vitro.

摘要

Cells in higher eukaryotes package genetic information into chromosomes. The chromosome itself has many levels of structure of its chromatin (nucleosomal array), consisting of the 10nm fiber, the 30nm fiber, and intermediates of the two. With the development of DNA sequences that strongly position octamer, in depth structural studies of the chromatin fiber have been possible. Here, analytical ultracentrifugation analysis follows the compaction of arrays with varying linker lengths. It has been shown that of the four arrays used, three (179-12, 199-12, 222-12) have sedimentation coefficients of 30-35S (15912 has an S-value of 46) which are very similar. XL-A analysis shows that the fibers possess intrinsic structural differences in the sedimentation coefficients of 30nm fibers: 159-12 is 55S, 179-12 is 43S, 199-12 is 50S, and 222-12 is 62S. To determine if accessibility of the histone octamer changes between the different arrays, crosslinking with APB at unique cysteines was performed. Crosslinking studies confirm that each array has different accessibility.