a, Heat map of marker genes of populations from Fig. 3a. Statistics for differential gene expression tests were applied to n = 13,185 cells. b, c, Heat map of differentially expressed genes between wild-type and Hand2-null OFT (b) and RV (c) cells captured at E7.75 and E8.25. Statistics were applied to n = 253 OFT cells per genotype at E7.75, n = 276 OFT cells per genotype at E8.25, n = 132 RV cells per genotype at E7.75, and n = 331 RV cells per genotype at E8.25. Scale indicates z-scored expression values. d, Quantification of fluorescence signal for indicated genes in f and Fig. 3h–j. n = 3 independent embryos per genotype. The mean ± s.e.m. is indicated. Two-tailed t-test. *P < 0.05 and **P < 0.01. e, Violin plots showing expression of Smyd1 and Sema3c in wild-type and Hand2-null E8.25 AHF cells. f, Expression of Hoxb1 in wild-type and Hand2-null embryos at E9.25 by whole-mount in situ hybridization in right lateral view. Arrowheads indicate expanded anterior Hoxb1 expression in Hand2-null embryos. Scale bars, 500 μm. n = 3 independent experiments with similar results. g, Proportion of wild-type and Hand2-null cells from each population captured at E7.75. n = 5 wild-type embryos and n = 3 Hand2-null embryos. The mean ± s.e.m. is indicated. Two-tailed t-test. *P < 0.05 and **P < 0.01. h, Violin plots showing expression of Nppa and Nppb in E8.25 wild-type and Hand2-null RV cells. All genes in a–c, e, h have a Bonferroni correction adjusted P < 1 × 10−4 (two-sided Wilcoxon rank-sum test). Violin plot summary statistics: centre white line represents median gene expression and central black rectangle spans the first to third quartile of the data distribution. The whiskers indicate values that are 1.5× the interquartile range above the third quartile or below the first quartile. Source data