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- Title
Droplet-based high-throughput single microbe RNA sequencing by smRandom-seq.
- Authors
Xu, Ziye; Wang, Yuting; Sheng, Kuanwei; Rosenthal, Raoul; Liu, Nan; Hua, Xiaoting; Zhang, Tianyu; Chen, Jiaye; Song, Mengdi; Lv, Yuexiao; Zhang, Shunji; Huang, Yingjuan; Wang, Zhaolun; Cao, Ting; Shen, Yifei; Jiang, Yan; Yu, Yunsong; Chen, Yu; Guo, Guoji; Yin, Peng
- Abstract
Bacteria colonize almost all parts of the human body and can differ significantly. However, the population level transcriptomics measurements can only describe the average bacteria population behaviors, ignoring the heterogeneity among bacteria. Here, we report a droplet-based high-throughput single-microbe RNA-seq assay (smRandom-seq), using random primers for in situ cDNA generation, droplets for single-microbe barcoding, and CRISPR-based rRNA depletion for mRNA enrichment. smRandom-seq showed a high species specificity (99%), a minor doublet rate (1.6%), a reduced rRNA percentage (32%), and a sensitive gene detection (a median of ~1000 genes per single E. coli). Furthermore, smRandom-seq successfully captured transcriptome changes of thousands of individual E. coli and discovered a few antibiotic resistant subpopulations displaying distinct gene expression patterns of SOS response and metabolic pathways in E. coli population upon antibiotic stress. smRandom-seq provides a high-throughput single-microbe transcriptome profiling tool that will facilitate future discoveries in microbial resistance, persistence, microbe-host interaction, and microbiome research. Population level transcriptomics measurements miss bacterial heterogeneity. Here the authors report smRandom-seq, a droplet-based high-throughput single-microbe RNA-seq assay, using random primers for in situ cDNA generation, droplets for single-microbe barcoding, and CRISPR-based rRNA depletion.
- Subjects
RNA sequencing; GENE expression; SPECIES specificity; CRISPRS; MICROORGANISMS; ESCHERICHIA coli
- Publication
Nature Communications, 2023, Vol 14, Issue 1, p1
- ISSN
2041-1723
- Publication type
Article
- DOI
10.1038/s41467-023-40137-9