{"database":"GEO","file_versions":[{"headers":{"Content-Type":["application/json"]},"body":{"files":{"Other":["ftp://ftp.ncbi.nlm.nih.gov/geo/series/GSE273nnn/GSE273521/"]},"type":"primary"},"statusCode":"OK","statusCodeValue":200}],"scores":null,"additional":{"omics_type":["Methylation profiling"],"species":["Mus musculus"],"gds_type":["Methylation profiling by high throughput sequencing"],"full_dataset_link":["https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE273521"],"repository":["GEO"],"entry_type":["GSE"],"additional_accession":[]},"is_claimable":false,"name":"A theoretical and experimental framework enables low-coverage sequencing for accurate quantification of genome-wide cytosine modification levels","description":"5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) regulate gene expression and exhibit dynamic levels during development and disease. While high-depth, base-resolution studies offer the most detailed view of epigenetic landscapes, many open questions are answered by surveying changes in 5mC/5hmC levels across larger cohorts. Nonetheless, current global quantification methods, including mass spectrometry, are typically limited in accessibility, accuracy, or throughput. Here, to evaluate the viability of low-coverage sequencing as an alternative, we first computationally downsampled deeply sequenced data to resolve the three-way relationship between sequencing coverage, modification levels, and measurement error. This relationship allowed us to develop a facile online tool for error calculation and to define experimental targets: <0.24% genome coverage can quantify 5mC and low-abundance 5hmC with minimal and predictable errors (<5%). Importantly, in direct comparisons, low-depth sequencing (Sparse-Seq) demonstrated high accuracy and less variability than mass spectrometry, while distinctively preserving genomic context. Applied serially to developing mouse brains, Sparse-Seq revealed an earlier emergence of 5hmCpG compared to 5mCpH and uncovered previously overlooked, genomic feature-specific epigenetic changes. This work establishes a rigorous foundation for employing Sparse-Seq as a highly accessible approach for 5mC/5hmC quantification, enabling economical first-pass analysis of epigenetic landscapes suited for large cohort studies and new hypothesis generation.","dates":{"publication":"2026/06/21"},"accession":"GSE273521","cross_references":{"GSM":["GSM8431616","GSM8431585","GSM8431584","GSM8431583","GSM8431582","GSM8431581","GSM8431580","GSM8431604","GSM8431603","GSM8431569","GSM8431602","GSM8431601","GSM8431589","GSM8431600","GSM8431588","GSM8431587","GSM8431586","GSM8431609","GSM8431608","GSM8431607","GSM8431606","GSM8431605","GSM8431596","GSM8431574","GSM8431595","GSM8431573","GSM8431572","GSM8431594","GSM8431593","GSM8431571","GSM8431592","GSM8431570","GSM8431591","GSM8431590","GSM8431615","GSM8431614","GSM8431613","GSM8431612","GSM8431579","GSM8431578","GSM8431611","GSM8431599","GSM8431577","GSM8431610","GSM8431598","GSM8431576","GSM8431575","GSM8431597"],"GPL":["16417"],"GSE":["273521"],"taxon":["Mus musculus"]}}