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Here we present DUET\u2010seq, an open\u2010source droplet microfluidic platform for joint profiling of chromatin accessibility and gene expression from the same nucleus. DUET\u2010seq combines programmable dissolvable dual\u2010linker hydrogel beads with one\u2010step intra\u2010droplet RT\u2010PCR to physically co\u2010index RNA and transposed chromatin fragments, completing library preparation within 12 h. Optimization of joint reaction conditions, including suppression of residual Tn5 activity, achieves \u223c3,200 genes per HEK293T cell while maintaining high\u2010quality chromatin accessibility profiles. By providing a fully disclosed reagent\u2010and\u2010device ecosystem, DUET\u2010seq enables researchers to optimize lysis and reaction chemistry for non\u2010standard tissues without proprietary constraints. We demonstrate the platform's versatility by mapping over 14,000 cis\u2010regulatory element\u2010to\u2010gene linkages in adult mouse brain and by profiling \u223c29,000 nuclei across six stages of postnatal spermatogenesis, where paired measurements reveal two modes of temporal regulatory decoupling\u2014epigenetic priming and chromatin inertia\u2014that are inaccessible to unimodal assays. DUET\u2010seq thus provides an accessible, cost\u2010effective framework for joint single\u2010nucleus multi\u2010omic profiling, with broad applicability across developmental biology, disease epigenomics, and functional genomics.<\/jats:p>","DOI":"10.1002\/advs.77986","type":"journal-article","created":{"date-parts":[[2026,9,27]],"date-time":"2026-09-27T18:00:31Z","timestamp":1790532031000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["DUET\u2010seq: An Open\u2010Source Droplet Platform for High\u2010Fidelity Joint Chromatin and Transcriptome Profiling Reveals Temporal Regulatory Decoupling in Single Cells"],"prefix":"10.1002","author":[{"given":"Dong","family":"Cheng","sequence":"first","affiliation":[{"name":"Department of Infectious Diseases Key Laboratory of Molecular Biology For Infectious Diseases (Ministry of Education) Institute For Viral Hepatitis The Second Affiliated Hospital Chongqing Medical 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