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. 2017 Mar 1;6(5):383–392. doi: 10.1016/j.molmet.2017.02.007

Figure 1.

Figure 1

FACS sorting and single-cell RNA sequencing of POMC-eGFP neurons. (A) The FACS gating for GFP (530 nm) and reference channel (460 nm). The GFP positive cells from the mediobasal hypothalamus (MBH Pos) are labelled blue, and negative cells (MBH Neg) are collected from the orange square within the negative population (purple). The cells were further gated for nuclear stain (DraQ5), cell size (FSC), and granularity (SSC), data not shown. (B) t-SNE was used to show the segregation of cells (MBH Pos and Neg, as well as GFP positive cells collected from the pituitary) based on their transcriptomic profiles. (C) Quality checks for FACS showing the GFP intensity and cell size measurement among the 3 cell types. We also examined for RNA expression of Gfp mRNA, neuronal marker Ncam1, the housekeeping genes Actb and Gapdh, Pomc, and Cartpt. (D)Pcsk1 and Pcsk2 are widely expressed in the cells sequenced. (E) We detected an average of 6882 genes per cell (≥5 RPM), adding all the genes from all the 163 cells together resulted in a total of 12,187 detected genes, indicating heterogeneity in gene expression.