title: Enzymatic and chemical mapping of nucleosome distribution in purified micro- and macronuclei of the ciliated model organism, Tetrahymena thermophila.
abstract: Genomic distribution of the nucleosome, the basic unit of chromatin, contains important epigenetic information. To map nucleosome distribution in structurally and functionally differentiated micronucleus (MIC) and macronucleus (MAC) of the ciliate Tetrahymena thermophila, we have purified MIC and MAC and performed micrococcal nuclease (MNase) digestion as well as hydroxyl radical cleavage. Different factors that may affect MNase digestion were examined, to optimize mono-nucleosome production. Mono-nucleosome purity was further improved by ultracentrifugation in a sucrose gradient. As MNase concentration increased, nucleosomal DNA sizes in MIC and MAC converged on 147 bp, as expected for the nucleosome core particle. Both MNase digestion and hydroxyl radical cleavage consistently showed a nucleosome repeat length of ~200 bp in MAC of Tetrahymena, supporting ~50 bp of linker DNA. Our work has systematically tested methods currently available for mapping nucleosome distribution in Tetrahymena, and provided a solid foundation for future epigenetic studies in this ciliated model organism.
authors: Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, 266003, China.
authors: Xiao
authors: X
authors: Chen
authors: Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, 266003, China. shangao@ouc.edu.cn.|Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266003, China. shangao@ouc.edu.cn.
authors: Shan
authors: S
authors: Gao
authors: Department of Pathology, University of Michigan, Ann Arbor, MI, 48109, USA.
authors: Yifan
authors: Y
authors: Liu
authors: Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, 266003, China.
authors: Yuanyuan
authors: Y
authors: Wang
authors: Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, 266003, China.
authors: Yurui
authors: Y
authors: Wang
authors: Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, 266003, China. wsong@ouc.edu.cn.
authors: Weibo
authors: W
authors: Song
mesh_terms: D002499:Centrifugation, Density Gradient; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D016798:Ciliophora / Q000235:genetics* / Q000378:metabolism; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004587:Electrophoresis, Agar Gel; D048648:Macronucleus / Q000235:genetics* / Q000378:metabolism; D008836:Micrococcal Nuclease / Q000378:metabolism; D009707:Nucleosomes / Q000235:genetics* / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013395:Sucrose; D016808:Tetrahymena thermophila / Q000235:genetics* / Q000378:metabolism
chemical_list: D002843:Chromatin; D016054:DNA, Protozoan; D009707:Nucleosomes; D013395:Sucrose; D008836:Micrococcal Nuclease
keywords: Tetrahymena; hydroxyl radical cleavage; macronuclei; micrococcal nuclease digestion; micronuclei; nucleosome