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. Author manuscript; available in PMC: 2020 Nov 24.
Published in final edited form as: Cell Stem Cell. 2020 Sep 28;27(5):765–783.e14. doi: 10.1016/j.stem.2020.09.001

Figure 3: Heterozygous PRS enhancer deletion in vitro impacts SOX9 expression during a restricted window of development.

Figure 3:

(A) Overview of differentiation, including early hCNCCs at day 11 (D11), passage 1–2 early hCNCCs (P1–2), P3–4 late hCNCCs, and chondrocytes at day 5 and 9.

(B) Schematic of allele-specific RT-ddPCR indicating primers and LNA probes (HEX/FAM) for T/C SNP (rs74999341) in SOX9 3’UTR. Wildtype (left), heterozygous EC1.45 deletion (right).

(C) RT-ddPCR for wildtype (green boxplot) and EC1.45 heterozygous deletion (red), plotting SOX9 C:T expression ratio.

(D) ATAC-seq reveals hCNCC-specific accessibility for EC1.45. Representative traces from 3–4 replicates.

(E) Luciferase assay for late hCNCCs (left) and chondrocytes (right). A COL2A1 enhancer is active in both cell types, while EC1.45 and EC1.25 become inactive in chondrocytes.

See also Figure S4 and Table S3.