MINICHROMOSOMES INTRODUCED IN CELL NUCLEI

细胞核中引入微型染色体

• 批准号: 3294095
• 负责人: MASARU RYOJI
• 金额: $ 13.35万
• 依托单位: SCRIPPS CLINIC AND RESEARCH FOUNDATION
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-05-01 至 1990-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3294095
• 关键词: cell nucleus; chromatin; chromosomes; circular DNA; conformation; egg /ovum; electron microscopy; electrophoresis; fresh water environment; gene expression; genetic manipulation; genetic mapping; genetic recombination; genetic transcription; microinjections; nucleic acid sequence; nucleosomes; radiotracer

Our goal is to understand how the structure of chromatin is involved in the regulation of gene expression and replication. Several specific questions will be asked by employing minichromosomes that are assembled on DNA injected in frog oocytes and eggs. Firstly, the structure of the peculiar "dynamic" minichromosomes will be studied. These minichromosomes carry unconstrained DNA supercoils, and are thought to represent transcriptionally competent chromatin. Dynamic minichromosomes will be separated from the conventional "static" minichromosomes, and their structure will be characterized by nuclease digestions, by electrophoretic identification of protein composition, and by electron microscopic observations. Taking advantage of the small size of the 5S RNA gene, further efforts will directed to isolate and analyze a fraction of dynamic minichromosomes that are actually engaged in transcription. Secondly, replication of dynamic minichromosomes will be studied with respect to whether they are preferentially replicated relative to the conventional, static minichromosomes, and whether the dynamic structure can be directly propagated to the progeny minichromosomes. The efficient assembly of dynamic minichromosomes on the preformed complex of 5S DNA and transcription factor TFIIIA will be exploited to examine such a possibility. Thirdly, nucleosome positions in inactive, static minichromosomes carrying a gene for either 5S RNA or histone H4 will be examined in order to test the hypothesis that the gene inactivation could be a result of blockage of promoter sequences by a nucleosome core. Minichromosomes carrying 32P label at a unique restriction site will be used to map nucleosome positions.

我们的目标是了解染色质的结构是如何参与到