{"database":"panorama","file_versions":[],"scores":null,"additional":{"omics_type":["Proteomics"],"submitter":["Nobuaki Takemori"],"species":["Drosophila Melanogaster"],"full_dataset_link":["https://panoramaweb.org/3DJiW8.url"],"submitter_email":["takemori@m.ehime-u.ac.jp"],"submitter_affiliation":["Ehime University"],"sample_protocol":[""],"repository":["PanoramaPublic"],"data_protocol":[""],"pubmed_abstract":["DNA is packaged with histones to form chromatin that impinges on all nuclear processes, including transcription, replication and repair, in the eukaryotic nucleus. A complete understanding of these molecular processes requires analysis of chromatin context in vitro. Here, Drosophila four core histones were produced in a native and unmodified form using wheat germ cell-free protein synthesis. In the assembly reaction, four unpurified core histones and three chromatin assembly factors (dNAP-1, dAcf1 and dISWI) were incubated with template DNA. We then assessed stoichiometry with the histones, nucleosome arrays, supercoiling and the ability of the chromatin to serve as a substrate for histone-modifying enzymes. Overall, our method provides a new avenue to produce chromatin that can be useful in a wide range of chromatin research."],"pubmed_title":["De novo reconstitution of chromatin using wheat germ cell-free protein synthesis."],"pubmed_authors":["Endo Yaeta Y, Takemori Nobuaki N, Nagy Szilvia K SK, Okimune Kei-Ichi KI, Kamakaka Rohinton R, Onouchi Hitoshi H, Takasuka Taichi E TE"],"additional_accession":[]},"is_claimable":false,"name":"De novo reconstitution and isolation of nucleosomes using the wheat germ cell-free protein synthesis","description":"DNA is packaged with histones and non-histone proteins to form chromatin that impinges on all nuclear processes such as transcription, replication and repair. A complete molecular understanding of these processes requires analysis of chromatin context. In this study, we report a simple and robust system using the wheat cell-free protein synthesis to assemble chromatin. We report two assembly protocols, the post-translational nucleosome assembly (PNAP) and co-translational nucleosome assembly (CNAP), both of which resulted in a native and unmodified form of chromatin. In the PNAP, a mixture of four unpurified core histones and chromatin assembly factors was combined with other components and incubated with DNA. In the CNAP, mRNAs encoding for these proteins were incubated with DNA in the translation mixture, and concomitantly assembled chromatin. Furthermore, we developed a rapid one-step method to isolate the assembled chromatin using divalent cations. Isolated chromatin was assessed for the stoichiometry of histones, nucleosome arrays, supercoiling, and as a substrate for histone modification enzymes. Our methods provide a new avenue to assemble customized chromatin to understand chromatin structure and function.","dates":{"publication":"Wed Jun 17 00:00:00 BST 2026"},"accession":"PXD011113","cross_references":{"TAXONOMY":["7227"],"pubmed":["33960726"]}}