Fun30 regulates chromatin structure at the HMR boundary element Chromatin analysis using micrococcal nuclease digestion and indirect endlabeling.
Spheroplasts were generated from both a wild-type yeast strain and the isogenic fun30 mutant strain and incubated briefly in the presence of increasing concentrations of micrococcal nuclease. The purified genomic DNA was then separated on an agarose gel and analyzed after ethidium bromide staining (left panels) or by Southern blot analysis for the HMR boundary region via indirect end-labelling (right panels). Asterisks indicate major digestion products. The position of the tRNA barrier is indicated by a small box. The arrow indicates the direction of transcription of the tRNA. The coding regions for a1 and GIT1 lie upstream and downstream, respectively, of the analysed fragment. The molecular size marker (M) is in multiples of 100 bp.