The research paper from the Kato Lab has been published in the Gene!
A low-input CUT&RUN-qPCR method for histone modification profiling in Daphnia magna embryos
Nikko Adhitama, Daffa Ramadhan Aridis, Nong Dang Quang, Pijar Religia, Yasuhiko Kato, Hajime Watanabe
Abstract
Understanding how organisms translate environmental variation into phenotypic variation is a central question in ecology and evolution. Histone modifications are key epigenetic regulators underlying phenotypic plasticity, enabling the generation of distinct phenotypes from a single genome. Cleavage Under Targets and Release Using Nuclease (CUT&RUN) is a powerful method for profiling histone modifications from low amounts of starting material. Here, we adapted a low-input CUT&RUN-qPCR protocol for embryos of the freshwater crustacean Daphnia magna, an emerging model for studying the molecular mechanisms of phenotypic plasticity. We established a cell dissociation protocol and applied the method to the male-determining gene doublesex1 (dsx1) locus during environmentally induced sex determination. CUT&RUN-qPCR detected male-biased enrichment of the active histone mark H3K4me3 at the dsx1 transcription start sites, whereas the repressive marks H3K9me3 and H3K27me3 showed higher enrichment in females. Using a dsx1 reporter strain in which dsx1-expressing cells are labeled with mCherry, we estimated that mCherry-positive cells comprise approximately 11% of dissociated male embryonic cells. Despite this heterogeneity, sex-biased H3K4me3 enrichment at dsx1 remained detectable from cells derived from as few as 20 embryos, corresponding to on the order of a few thousand target cells per reaction. This benchmark provides a practical guideline for planning CUT&RUN input requirements in heterogeneous and low-input samples. Our study establishes CUT&RUN-qPCR as a practical approach for histone modification profiling in Daphnia and provides a methodological basis for future work on the epigenetic underpinnings of phenotypic plasticity.
Gene, 1010, 150317. doi: 10.1016/j.gene.2026.150317. (2026)
https://www.sciencedirect.com/science/article/pii/S0378111926003276
