Supplementary Information for
Lesch BJ, Dokshin GA, Young RA, McCarrey JR, Page DC.
A set of genes critical to development is epigenetically poised in mouse germ cells from fetal stages through completion of meiosis.
PNAS 110, 16061 (2013)
- Supplementary Table 1 Log2fold-change values for differentially expressed genes (P=0.05) in fetal germ cells (index for Fig. S1)
- Supplementary Table 2 Pearson correlations between biological replicates for mean signal in nonoverlapping 5-kb genomic intervals
- Supplementary Table 3 Top five enriched GO categories enriched in genes associated with H3K4me3 or H3K27me3, using pooled replicates from onerepresentative sex and time point (female, E13.5)
- Supplementary Table 4 Mann–WhitneyPvalues for differential H3K4me3 or H3K27me3 signal in the“meiosis”and“cellular response to retinoic acid”GO categories, comparing sexes and time points
- Supplementary Table 5 Genes identified as poised in the mouse germ line
- Figure S1 Heat map showing significant changes in gene expression between male and female germ cells at each developmental time point (Topthree rows,male divided by female), between time points in female germ cells (Middlethree rows, later divided by earlier time points), or between time points in malegerm cells (Bottomthree rows, later divided by earlier time points). Genes were included if they were differentially regulated in at least one comparison atP=0.05 after correction for multiple hypothesis testing (two-sided analysis of variance using DESeq).
- Figure S2 Validation of small-scale ChIP-seq protocol using mouse embryonic stem cells (mESCs). (A) Pearson correlations between datasets for H3K4me3or H3K27me3 ChIP datasets. (Left) ChIP performed on 2×104vs. 1×105cells grown in the same dish. (Right) ChIP performed on 2×104cells vs.~1×108cells;1×108cell dataset from Mikkelsen et al. (1). (B) Sample gene tracks for data from 2×104,1×105,or1×108cells at the HoxB cluster (Left) and at a gene-richregion on chromosome 19 (chr19:3,922,435-4,515,456) (Right). (C) Overlap between genes associated with H3K4me3 or H3K27me3 in ChIP-seq from 2×104,1×105,or~1×108cells. (D) Pearson correlation between biological replicates in fetal germ cells. Shown are scatter-plots for median signal in 50-kb windowsfor replicates from female E12.5 germ cells, where ChIP was performed on different days, and female E12.5 compared with female E13.5 germ cells where ChIPwas performed on the same day.
- Figure S3 Boxplots showing the distribution of ChIP signal for each quartile of genes, based on expression level. The fourth quartile refers to the most highlyexpressed genes. Top and bottom of each box represents the 75th and 25th percentiles of ChIP signal, respectively; center line represents the median.Verticallines extend beyond the box up to 1.5-times the difference between the 75th and 25th percentiles. Hatchmarks represent outliers. For each expressionlevelquartile, the distribution of ChIP signal is statistically significantly different from that of the other quartiles (P=10-15, independent two-group Mann–WhitneyUtest).
- Figure S4 Expression, K4 signal, and K27 signal across fetal time points for genes with significantly changed expression in the“meiosis”GO category, and for allgenes in the“cellular response to RA”(RA, retinoic acid) GO category. Faint lines represent individual genes, and dark lines represent the mean. Red, female;blue, male.