Isolation of Histone from Sorghum Leaf Tissue for Top Down Mass Spectrometry Profiling of Potential Epigenetic Markers

Histones belong to a family of highly conserved proteins in eukaryotes. They pack DNA into nucleosomes as functional units chromatin. Post-translational modifications (PTMs) histones, which are dynamic and can be added or removed by enzymes, play critical roles regulating gene expression. In plants, epigenetic factors, including histone PTMs, related their adaptive responses the environment. Understanding molecular mechanisms control bring unprecedented opportunities for innovative bioengineering solutions. Herein, we describe protocol isolate nuclei purify histones from sorghum leaf tissue. The extracted analyzed intact forms top-down mass spectrometry (MS) coupled with online reversed-phase (RP) liquid chromatography (LC). Combinations stoichiometry multiple PTMs on same proteoform readily identified. addition, tail clipping detected using LC-MS workflow, thus, yielding global PTM profile core (H4, H2A, H2B, H3). We have applied this previously tissue collected large-scale field study, aimed at identifying markers drought resistance. could potentially adapted optimized chromatin immunoprecipitation-sequencing (ChIP-seq), studying similar plants.