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However, current high\u2010throughput mscRNA\u2010seq methods are limited by multiple centrifugation steps, which can lead to microbial loss and bias. smGel\u2010seq is reported, a high\u2010throughput single\u2010microbe RNA sequencing method for clinical microbiome samples that employs hydrogel beads to encapsulate individual microbes to reduce microbial loss and input requirements. In this method, a novel microchannel array device is implemented for encapsulating single microbe in dissolvable hydrogel beads (smDHBs), along with an optimized automated microfluidic platform to co\u2010encapsulate barcoded beads and smDHBs, enabling high\u2010throughput barcoding of individual microbes. smGel\u2010seq significantly increases the microbial recovery rate in a gut microbiome sample from 8.8% to 91.8%. Furthermore, this method successfully processes clinical microbiome samples with microbial inputs 20 times lower than those required by previous methods. Notably, smGel\u2010seq enables the first mscRNA\u2010seq in a clinical sputum microbiome sample, revealing a specific microbial subpopulation that may play a key role in environmental adaptability, antibiotic resistance, and pathogenicity. These results highlight the compatibility of smGel\u2010seq with clinical microbiome samples and demonstrate its potential for widespread application in diverse clinical and research settings.<\/jats:p>","DOI":"10.1002\/adma.202500481","type":"journal-article","created":{"date-parts":[[2025,4,9]],"date-time":"2025-04-09T02:34:30Z","timestamp":1744166070000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Fast Encapsulation of Microbes into Dissolvable Hydrogel Beads Enables High\u2010Throughput Microbial Single\u2010Cell RNA Sequencing of Clinical Microbiome Samples"],"prefix":"10.1002","volume":"37","author":[{"given":"Yuting","family":"Wang","sequence":"first","affiliation":[{"name":"Department of Laboratory Medicine of The First Affiliated Hospital &amp; Liangzhu Laboratory Zhejiang University School of Medicine  Hangzhou 311121 China"},{"name":"College of Biomedical Engineering and Instrument Science Zhejiang 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