Visualization of G4 Tetraplex DNA on Yeast Meiotic Chromosomes

9510732吉尔伯特本研究的目的是获得体内G4四链体DNA的证据;更具体地说，在减数分裂配对的染色体上。 生物化学和遗传学证据表明，这种新形式的DNA存在于体内，作为几个细胞过程的结构中间体，包括减数分裂，端粒功能和富含G的序列介导的重组。 酵母基因凯米编码G4-DNA依赖性核酸酶。 该基因的纯合缺失阻断了减数分裂I的酵母细胞，在减数分裂I中同源染色体配对但未能解析，支持G4-DNA参与减数分裂染色体配对的假设。 从减数分裂阻断的（kem 1）酵母细胞中分离裸染色体，通过光学显微镜、CHEF凝胶电泳和Southern印迹法观察和检查其DNA，并与野生型细胞的染色体进行比较。 来自keml（在减数分裂I中阻断的突变细胞）的染色体应该表现出更高级的结构变化，包括CHEF凝胶中的延迟的迁移率，以及来自同源染色体的DNA在多个点处连接的四聚体（4 N）复合物。 这项研究将揭示新的光减数分裂染色体配对，通过展示DNA-DNA相互作用与G4-DNA配对在体内一致，并将建立基础，调查其他细胞过程中，这种四链体DNA可能参与，如端粒功能和重组。 本研究的目的是为减数分裂配对染色体中新的G4四链体DNA结构提供证据。 生物化学和遗传学证据表明，这种新形式的DNA存在于体内，作为几个细胞过程的结构中间体，包括减数分裂，端粒功能和富含G的序列介导的重组。 ***

9510732 Gilbert The objective of this research is to obtain evidence for G4 tetraplex DNA in vivo; more specifically, on meiotically paired chromosomes. Biochemical and genetic evidence suggests that this novel form of DNA exists in vivo as a structural intermediate for several cellular processes including meiosis, telomere functioning, and G-rich sequence mediated recombination. The yeast gene KEMI encodes a G4-DNA dependent nuclease. A homozygous deletion of this gene blocks yeast cells at meiosis I, where homologous chromosomes are paired but fail to resolve, supporting the hypothesis that G4-DNA is involved in meiotic chromosome pairing. Naked chromosomes will be isolated from meiotically blocked (kem1() yeast cells and their DNA will be visualized and examined by a light microscopy, and by CHEF gel electrophoresis and Southern blotting in comparison with chromosomes from wild type cells. Chromosomes from keml( mutant cells blocked in meiosis I should exhibit higher-order structural changes including retarded mobility in CHEF gels, and tetrameric (4N) complexes of DNA from homologous chromosomes joined at multiple points. This research will shed new light on meiotic chromosome pairing by demonstrating DNA-DNA interaction consistent with G4-DNA pairing in vivo, and will establish the foundation to investigate other cellular processes in which this tetraplex DNA might be involved, such as telomere functioning and recombination. %%% The objective of this research is to obtain evidence for the novel G4 tetraplex DNA structure in meiotically paired chromosomes. Biochemical and genetic evidence suggests that this novel form of DNA exists in vivo as a structural intermediate for several cellular processes including meiosis, telomere functioning, and G-rich sequence mediated recombination. ***