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. 2022 Aug 21;9(29):2201273. doi: 10.1002/advs.202201273

Figure 1.

Figure 1

Osteoblast selection in single cell RNA sequencing (scRNASeq) based on marker genes. A) Left: individual mRNA counts based on unique molecular identifiers (UMI) for Col1a1, Runx2, Sp7, and Ibsp in cell isolates from femurs and tibias of E18.5 embryos (3 Hom, 511 osteoblasts (OBs); 3 Het, 96 OBs; 4 WT), 485 OBs). Right: UMIs in OBs selected based on the indicated threshold values (optimized as described in Experimental Section). Runx2 and Sp7 transcripts are not detected by scRNASeq in some OBs expressing these genes because of their relatively low abundance. Ibsp is also not detected in some OBs because of its highly variable expression dependent on OB differentiation. B) Validation of OB marker gene selection by mRNA‐FISH in 5 µm paraffin sections of E18.5 femurs. Co‐expression of Col1a1 and Sp7 and/or Col1a1 and Ibsp is observed in all OBs (bone surface cells producing collagen I) and no other cells. White boxes indicate zoomed areas. Yellow arrows point to osteoblasts expressing Col1a1 and Sp7 (left) or Col1a1 and Ibsp (right). Green arrows point to periosteal fibroblasts expressing only Col1a1. Red arrows point to chondrocytes expressing only Sp7 or Ibsp.