ISDH-seq: a robust methodology for profiling and characterization of open chromatin

We present in situ DNase I hypersensitivity sequencing (ISDH-seq), a novel methodology for genome-wide open chromatin characterization in plants, which can be applicable to 100-200K nuclei. Compared to conventional DNase-seq and ATAC-seq applied to the same rice tissues, ISDH-seq detects 72% and 120% more open chromatin sites (OCSs), respectively. ISDH-specific OCSs exhibit unique epigenetic signatures: hypomethylation, co-enrichment with H3K27me3, and strong spatial chromatin interaction associations.ISDH-seq overcomes method-specific biases in OCS detection, expands the plant regulatory genome atlas, and offers cross-species applicability. This technology provides critical insights into noncoding regulatory mechanisms and their biological implications in plant genomes. This study profiles open chromatin landscapes in Oryza sativa (rice) using two-week-old seedlings under untreated conditions. We developed ISDH-seq (in situ DNase I hypersensitivity sequencing) and compared its performance with DNase-seq and ATAC-seq on the same batch of seedling tissues (Biological replicates n=2).

本研究报道了原位DNase I超敏测序（in situ DNase I hypersensitivity sequencing，ISDH-seq），这是一种用于植物全基因组开放染色质表征的新型方法，可适配10万至20万个细胞核。相较于针对同一水稻组织的传统DNase-seq与ATAC-seq，ISDH-seq分别多检测到72%和120%的开放染色质位点（open chromatin sites，OCSs）。ISDH-seq特有的OCSs呈现独特的表观遗传特征：低甲基化、与H3K27me3共富集，以及显著的空间染色质互作关联。ISDH-seq克服了OCS检测中方法特异性的偏差，拓展了植物调控基因组图谱，并具备跨物种应用潜力。该技术为解析植物基因组中非编码调控机制及其生物学意义提供了关键视角。本研究以未处理条件下生长两周的水稻（Oryza sativa）幼苗为材料，绘制了其开放染色质图谱。我们开发了ISDH-seq（原位DNase I超敏测序），并以同一批次的幼苗组织（生物学重复n=2）为样本，对比了其与DNase-seq、ATAC-seq的检测性能。