PRJNA328774: A single-cell transcriptomic map of the human and mouse pancreas reveals inter- and intra-cell population structure
Summary: While the function of the mammalian pancreas hinges on complex interactions of distinct cell types, gene expression profiles have primarily been described with bulk mixtures of cells. Here, we invoked inDrop, a droplet-based single-cell RNA-Seq method, to determine the transcriptomes of over 12,000 individual pancreatic cells from four human donors and two strains of mice. Cells could be divided into 15 clusters that matched previously characterized cell types: all endocrine cell types, including rare ghrelin-expressing epsilon-cells, exocrine cell types, vascular cells, Schwann cells, quiescent and activated pancreatic stellate cells, and four types of immune cells. We detected subpopulations of ductal cells with distinct expression profiles, and validated their existence with immuno-histochemistry stains. Moreover, among human beta-cells, we detected heterogeneity in the regulation of genes relating to functional maturation and levels of ER-stress. Finally, we deconvolve bulk gene expression samples using single-cell RNA-Seq to detect disease associated differential expression. Thus, our cross-species dataset provides a resource for the discovery of novel cell type-specific and cell type-restricted transcription factors, signaling receptors, and medically-relevant genes.
Overall Design: Single-cell RNA sequencing of pancreatic islets from 4 human donors and 2 mice strains.
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Human pancreatic islets were obtained from Prodo or NDRI and recovered in CMRLS at 37°C for 24h-48h hours after receipt. Mouse islets were prepared using gradient centrifugation and islets from the same strain were pooled, then recovered in XX for 24h. |
| Extract Protocol: |
Cells were encapsulated using the inDrop platform, into droplets on ice and lysed in the 4nL microfluidic droplets using a final concentration of 0.4% NP-40. Single cell lysates were subject to reverse transcription at 50°C without purification of RNA (as previously described in Klein et al., Cell 2015).Cells were barcoded using the inDrop platform (Klein et al., Cell 2015), which makes use of the CEL-Seq protocol for library construction Hashimshony et al., Cell Reports 2012) |
| Library Construction Protocol: |
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poly(A)+ RNA |
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PAIRED |
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Forward |
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ILLUMINA |
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Illumina HiSeq 2500 |
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Specific |
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Spike-In |
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A Single-Cell Transcriptomic Map of the Human and Mouse Pancreas Reveals Inter- and Intra-cell Population Structure.
Cell systems . 2016-09-22 [PMID:
27667365]
