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. 2011 Aug 18;286(41):35347–35357. doi: 10.1074/jbc.M111.281923

FIGURE 1.

FIGURE 1.

Changes of histone H1 variant gene expression during the differentiation of human embryonic stem cells. A, silencing of NANOG, SOX2, and OCT4 genes during the differentiation of ES[4] stem cells. ES[4] cells were cultured in differentiation media as described under “Experimental Procedures” for 4, 8, 15, or 20 days. At the indicated time points, the cells were processed for RNA extraction. Expression of NANOG, SOX2, and OCT4 genes was determined by RT-qPCR with specific oligonucleotide pairs. GAPDH expression was measured for normalization, and the data are shown relative to the maximal level of each gene. B, comparison of H1 variant gene expression in ES cells. RNA extracted from ES[4] cells was used to measure histone H1 expression by RT-qPCR with variant-specific oligonucleotide pairs, as well as FOXA2 and SOX2 expression with oligonucleotide pairs in the same exon. GAPDH was used for normalization. To allow comparison between genes, each cDNA value was normalized to the value of qPCR amplification of genomic DNA (gDNA) extracted from ES[4], with the same primer set. C, histone H1 variant gene expression during the differentiation of ES[4]. The RNA samples of the ES[4] differentiation curve described in A were used to measure histone H1 expression by RT-qPCR with variant-specific oligonucleotide pairs. GAPDH was used for normalization. To allow comparison between H1 variants, each histone H1 cDNA value was normalized to the value of qPCR amplification of genomic DNA as in B. D, proportion of histone H1 variant transcripts in differentiated and undifferentiated cells. The relative proportion of each H1 variant expression in the initial and final time points along the differentiation kinetic was calculated from the data in C and expressed as percentage of the total H1 content. In addition to ES[4], data for ES[2] differentiation are also shown calculated from a kinetic shown in supplemental Fig. S1. For the ES[2] experiment, day 4 is the first time point available. The means and S.D. of a representative experiment quantified in triplicate are shown.