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Droplet-based high-throughput single microbe RNA sequencing by smRandom-seq.

Ziye XuYuting WangKuanwei ShengRaoul RosenthalNan LiuXiaoting HuaTianyu ZhangJiaye ChenMengdi SongYuexiao LvShunji ZhangYingjuan HuangZhaolun WangTing CaoYifei ShenYan JiangYunsong YuYu ChenGuoji GuoPeng YinDavid A WeitzYongcheng Wang
Published in: Nature communications (2023)
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.
Keyphrases
  • single cell
  • rna seq
  • high throughput
  • gene expression
  • genome wide
  • escherichia coli
  • endothelial cells
  • dna methylation
  • crispr cas
  • microbial community
  • quantum dots