De novo deposition of the histone H3 variants H3.1 and H3.3 during DNA replication
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ABSTRACT: We developed a new sequencing assay to track the de novo deposition of the histone H3 variants H3.1 and H3.3 during S phase. We use cells stably expressing H3.1-SNAP or H3.3-SNAP, and synchronize them in G1/S by double-thymidine block. The SNAP-tag enables to discriminate newly synthesized histones from preexisting ones, via a quench-chase-capture strategy. We applied this strategy to isolate new H3.1 and H3.3 after releasing cells into S phase, and probed their distribution by MNase digestion and sequencing. We could thus characterize H3.1 and H3.3 dynamics from early to mid S phase at genome-wide resolution. We further applied our method to investigate the consequences of perturbations upon deletion of the H3.3 chaperone HIRA. We used HIRA knockout and control cells, and compared H3.1 and H3.3 distribution to early replication patterns by EdU labeling and sequencing of nascent DNA.
INSTRUMENT(S): Illumina HiSeq 2500, Illumina NovaSeq 6000
ORGANISM(S): Homo sapiens
SUBMITTER: Alberto Gatto
PROVIDER: E-MTAB-10619 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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