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Differences are demonstrated between A (transcriptionally active) and B (transcriptionally inactive) chromosomes that are characterized by a different level of histone H4 acetylation and a different timing of DNA replication. These differences between the chromatin of A and B chromosomes were found after immunolabelling of chromsomes of Brachycome dichromosomatica with antibodies specific for different acetylated forms (lysine 5, 8, 12 and 16) of histone H4. In contrast to the A chromosomes, which are labelled brightly in their entirety, the transcriptionally inactive B chromosomes are faintly labelled with antibodies against H4Ac5 and H4Ac8. No such difference between the chromosomes is found after immunostaining with the other antibodies H4Ac12 and H4Ac16. Analysis of DNA replication timing in root-tip meristems suggests that B chromosomes are labelled late in S-phase compared with A chromosomes. After C-banding the B chromosome appeared to have a similar amount of heterochromatin to the A chromosomes.