Map active enhancer and gene promoter regions in the SCN

Histone ChIP-seq was performed at varied time of the day to map the gene promoter and active enhancer sites in the central clock; SCN (suprachiasmatic nuclei). Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for the histone modifications H3K4me3 and H3K27ac in adult mouse SCN and cortex at distinct time of the day . Overall design: WT C57Bl/6J mice were used for SCN tissue collection at 6 distinct time-points starting from ZT3 at every 4 hours, where lights on at 7 am (ZT0) and lights off at 7 pm (ZT12). We also collected cortical punches at ZT3 and ZT15 to compare and establish SCN enriched chromatin modifications. For histone chromatin immunoprecipitation two separate biological replicates per time-point per tissue-type was collected where each biological replicate composed of 3-4 individual SCN or cortical punches.

本研究于一日内不同时间点开展组蛋白染色质免疫共沉淀测序（Histone ChIP-seq），以定位中枢生物钟核心——视交叉上核（suprachiasmatic nuclei, SCN）内的基因启动子与活性增强子位点。针对成年小鼠视交叉上核与大脑皮层的组蛋白修饰H3K4me3和H3K27ac，本研究同步采用染色质免疫共沉淀测序（Chromatin immunoprecipitation DNA-sequencing, ChIP-seq）进行检测，采样覆盖一日内多个不同时间点。

实验设计概况如下：选用野生型（wild type, WT）C57Bl/6J小鼠，以光照起始时间为上午7点（节律时间0点，Zeitgeber Time, ZT0）、光照结束时间为下午7点（节律时间12点，ZT12）的光照周期为基准，在节律时间3点（ZT3）起每间隔4小时采样，共设置6个不同时间点用于收集视交叉上核组织。此外，本研究还在ZT3与ZT15两个时间点采集大脑皮层穿刺样本，用于对比分析并确定视交叉上核富集的染色质修饰特征。针对组蛋白染色质免疫共沉淀实验，我们为每类组织的每个时间点设置2个独立生物学重复，每个生物学重复由3-4份独立的视交叉上核或大脑皮层穿刺样本组成。