Quartz-Seq: a highly reproducible and sensitive single-cell RNA sequencing method, reveals non-genetic gene-expression heterogeneity

AbstractDespite longstanding appreciation of gene expression heterogeneity in isogenic bacterial populations, affordable and scalable technologies for studying single bacterial cells have been limited. While single-cell RNA sequencing (scRNA-seq) has revolutionized studies of transcriptional heterogeneity in diverse eukaryotic systems, application of scRNA-seq to prokaryotes has been hindered by their extremely low mRNA abundance, lack of mRNA polyadenylation, and thick cell walls. Here, we present Prokaryotic Expression-profiling by Tagging RNA In Situ and sequencing (PETRI-seq), a low-cost, high-throughput, prokaryotic scRNA-seq pipeline that overcomes these technical obstacles. PETRI-seq uses in situ combinatorial indexing to barcode transcripts from tens of thousands of cells in a single experiment. PETRI-seq captures single cell transcriptomes of Gram-negative and Gram-positive bacteria with high purity and low bias, with median capture rates >200 mRNAs/cell for exponentially growing E. coli. These characteristics enable robust discrimination of cell-states corresponding to different phases of growth. When applied to wild-type S. aureus, PETRI-seq revealed a rare sub-population of cells undergoing prophage induction. We anticipate broad utility of PETRI-seq in defining single-cell states and their dynamics in complex microbial communities.

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Bioinformatics analysis of the gene expression profile of retinal pigmental epithelial cells based in single-cell RNA sequencing in myopic mice

Archives of Medical Science ◽

10.5114/aoms/131835 ◽

2021 ◽

Vol 17 (2) ◽

pp. 574-577

Author(s):

Ya Mo ◽

Mu-Lin He ◽

Jia-Zhen Yu ◽

Xue-Jun Xie

Keyword(s):

Gene Expression ◽

Epithelial Cells ◽

Single Cell ◽

Rna Sequencing ◽

Gene Expression Profile ◽

Expression Profile ◽

Bioinformatics Analysis ◽

Single Cell Rna Sequencing

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Microbial single-cell RNA sequencing by split-pool barcoding

Science ◽

10.1126/science.aba5257 ◽

2020 ◽

Vol 371 (6531) ◽

pp. eaba5257 ◽

Cited By ~ 2

Author(s):

Anna Kuchina ◽

Leandra M. Brettner ◽

Luana Paleologu ◽

Charles M. Roco ◽

Alexander B. Rosenberg ◽

...

Keyword(s):

Gene Expression ◽

Single Cell ◽

Rna Sequencing ◽

High Throughput ◽

Single Cell Analysis ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Growth Stages ◽

High Throughput Analysis ◽

Single Cell Rna Sequencing

Single-cell RNA sequencing (scRNA-seq) has become an essential tool for characterizing gene expression in eukaryotes, but current methods are incompatible with bacteria. Here, we introduce microSPLiT (microbial split-pool ligation transcriptomics), a high-throughput scRNA-seq method for Gram-negative and Gram-positive bacteria that can resolve heterogeneous transcriptional states. We applied microSPLiT to >25,000 Bacillus subtilis cells sampled at different growth stages, creating an atlas of changes in metabolism and lifestyle. We retrieved detailed gene expression profiles associated with known, but rare, states such as competence and prophage induction and also identified unexpected gene expression states, including the heterogeneous activation of a niche metabolic pathway in a subpopulation of cells. MicroSPLiT paves the way to high-throughput analysis of gene expression in bacterial communities that are otherwise not amenable to single-cell analysis, such as natural microbiota.

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Single-Cell Transcriptome Analysis Reveals Dynamic Cell Populations and Differential Gene Expression Patterns in Control and Aneurysmal Human Aortic Tissue

Circulation ◽

10.1161/circulationaha.120.046528 ◽

2020 ◽

Vol 142 (14) ◽

pp. 1374-1388

Author(s):

Yanming Li ◽

Pingping Ren ◽

Ashley Dawson ◽

Hernan G. Vasquez ◽

Waleed Ageedi ◽

...

Keyword(s):

Gene Expression ◽

Single Cell ◽

Rna Sequencing ◽

Aortic Wall ◽

Genome Wide Association ◽

Aortic Tissue ◽

Sequencing Data ◽

Genome Wide ◽

Single Cell Rna Sequencing ◽

Differential Gene

Background: Ascending thoracic aortic aneurysm (ATAA) is caused by the progressive weakening and dilatation of the aortic wall and can lead to aortic dissection, rupture, and other life-threatening complications. To improve our understanding of ATAA pathogenesis, we aimed to comprehensively characterize the cellular composition of the ascending aortic wall and to identify molecular alterations in each cell population of human ATAA tissues. Methods: We performed single-cell RNA sequencing analysis of ascending aortic tissues from 11 study participants, including 8 patients with ATAA (4 women and 4 men) and 3 control subjects (2 women and 1 man). Cells extracted from aortic tissue were analyzed and categorized with single-cell RNA sequencing data to perform cluster identification. ATAA-related changes were then examined by comparing the proportions of each cell type and the gene expression profiles between ATAA and control tissues. We also examined which genes may be critical for ATAA by performing the integrative analysis of our single-cell RNA sequencing data with publicly available data from genome-wide association studies. Results: We identified 11 major cell types in human ascending aortic tissue; the high-resolution reclustering of these cells further divided them into 40 subtypes. Multiple subtypes were observed for smooth muscle cells, macrophages, and T lymphocytes, suggesting that these cells have multiple functional populations in the aortic wall. In general, ATAA tissues had fewer nonimmune cells and more immune cells, especially T lymphocytes, than control tissues did. Differential gene expression data suggested the presence of extensive mitochondrial dysfunction in ATAA tissues. In addition, integrative analysis of our single-cell RNA sequencing data with public genome-wide association study data and promoter capture Hi-C data suggested that the erythroblast transformation-specific related gene( ERG ) exerts an important role in maintaining normal aortic wall function. Conclusions: Our study provides a comprehensive evaluation of the cellular composition of the ascending aortic wall and reveals how the gene expression landscape is altered in human ATAA tissue. The information from this study makes important contributions to our understanding of ATAA formation and progression.

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