Figure 4

Soluble and chromatin-bound histone isolation reveals distinct patterns of H2A and H2A.X-F modification during early development. (A) Cartoons of the embryo stages that we collected (drawings Copyright 1994 from Normal Table of Xenopus Laevis (Daudin) by Faber et al. Reproduced by permission of Garland Science/Taylor & Francis LLC). (B) Embryo fractionation scheme: five embryos per stage were collected, lysed, and homogenized, centrifuged at 1,000 g and the supernatant containing soluble histones was removed. The pellet was washed in the lysis buffer and then sonicated. This material was used as the chromatin fraction. (C) Equivalent volume of total soluble protein from the staged embryo fractionation was immunoblotted for linker histones, core histones, and the conserved H2A/H2A.X-F/H4 modifications as shown. Total soluble protein is shown in the Coomassie stained gel at the bottom. The period of transcriptional repression post fertilization is indicated at the bottom. The migration positions of H2A.X-F, H2A, and H4 are indicated on the left. (D) Equivalent volume of total chromatin protein from the staged embryo fractionation was immunoblotted for linker histones, core histones, and the conserved H2A/H2A.X-F/H4 modifications as shown. Total chromatin protein is shown in the Coomassie stained gel at the bottom. The stained histone protein bands are annotated. The period of transcriptional repression post fertilization is indicated at the bottom. The migration positions of H2A.X-F, H2A, and H4 are indicated on the right.