细胞功能测定

The Anaphase Promoting Complex (APC) ubiquitin ligase is critical for multiple processes including cell cycle, development, meiosis, and senescence. The importance of regulation of the APC by substrate competitive (pseudosubstrate) inhibitors, such as Emi1 and BubR1, has recent ...

Reverse genetic methods, such as homologous gene targeting, have greatly contributed to our understanding of molecular pathways in mitosis, especially in yeast. The chicken B-lymphocyte line, DT40, represents a unique example among vertebrate somatic cells where homologous gene ...

Ubiquitination and protein degradation regulate cell cycle progression in all eukaryotes. During mitosis, ubiquitination by the Anaphase-Promoting Complex/Cyclosome (APC/C) triggers sister chromatid separation and mitotic exit. The APC/C is tightly regulated by phosph ...

The anaphase-promoting complex/cyclosome (APC/C), a large (20S) multisubunit E3 ligase, has an essential role to ubiquitylate numerous substrates at specific times during mitosis and G1 phase as well as in meiosis. The deregulation of the APC/C causes cell death or genomic instability, wh ...

Cell cycle transitions are controlled, in part, by ubiquitin-dependent proteolysis. In mitosis, the metaphase to anaphase transition is governed by an E3 ubiquitin ligase called the cyclosome or Anaphase-Promoting Complex (APC), and a WD40-repeat protein co-factor called Cdc20. In v ...

The targeted destruction of key regulators helps to drive the cell cycle. Here we describe a quantitative assay to measure destruction of different regulators in mitotic cells. This assay uses GFP-tagged substrates and time-lapse fluorescence microscopy of single cells to pinpoint the ...

The spindle checkpoint is a surveillance mechanism that ensures the fidelity of chromosome segregation by inhibiting anaphase onset until all chromosomes have established stable bipolar attachments. Here we describe a number of protocols that can be used to assay the ability of buddi ...

In this chapter we describe the preparation of early mitotic C. elegans embryos for the tomographic reconstruction of end-morphologies of spindle microtubules. Early embryos are prepared by high-pressure freezing and freeze-substitution for thin-layer embedding in Epon/Ara ...

Kinetochores are multiprotein machines that initiate mitotic checkpoint signaling and control chromosome movement through interactions with microtubules. Our lab has utilized Xenopus laevis frog egg extracts to investigate the requirements for kinetochore assembly a ...

For high-fidelity chromosome segregation, kinetochores must be properly captured by spindle microtubules, but the mechanisms of initial kinetochore capture have remained elusive. Observation of individual kinetochore–microtubule interaction has been difficult, be ...

Capillary electrochromatography (CEC) is a micro-separation technique that combines the advantages of capillary zone electrophoresis with those of high-performance liquid chromatography. Accordingly, it has attracted extensive attention over the last decade. Among the ...

The components of the aminoglycosides, e.g., gentamicin, sisomicin, netilmicin, kanamycin, amikacin, and tobramycin, and related impurities of these antibiotics can be separated by means of micellar electrokinetic chromatography (MEKC). Derivatization with o-phthaldia ...

Microemulsion electrokinetic chromatography (MEEKC) is an attractive capillary electrophoretic technique in which a microemulsion is used as carrier electrolyte. Analytes may partition between the aqueous phase of the microemulsion and its oil droplets, which act as a pseudo ...

This chapter describes the technology and selected applications in the field of protein analysis for free flow electrophorsis (FFE). FFE is a highly versatile technology for applications in this field as a result of its continuous processing of sample and the possibility of separating alm ...

This chapter reviews the theory and methodological developments of on-line concentration techniques for the determination of environmental pollutant samples, such as organic and inorganic compounds in capillary zone electrophoresis (CZE) and also in micellar electrokin ...

Microtubules are required throughout plant development for a wide variety of processes, and different strategies have been evolved to visualize them. This chapter summarizes the most effective of these methods and points out potential problems and pitfalls. We outline the freeze-sh ...

Preservation of Tetrahymena thermophila basal body ultrastructure for visualization by transmission electron microscopy is improved by a combination of high pressure freezing (HPF) and freeze substitution (FS). These methods also reliably retain the antigenicity of cellu ...

The growth and maintenance of almost all cilia and flagella are dependent on the proper functioning of the process of intraflagellar transport (IFT). This includes the primary cilia of most cells in humans that are in interphase or the Go phase of the cell cycle. The model system for the study of IFT is the fl ...

The cytoskeleton is a complex of detergent-insoluble components of the cytoplasm playing critical roles in cell motility, shape generation, and mechanical properties of a cell. Fibrillar polymers−actin filaments, microtubules, and intermediate filaments− are major constit ...

The budding yeast Saccharomyces cerevisiae has many advantages as a model system, but until recently, high-resolution microscopy was not often attempted in this organism. Its small size, rounded shape, and rigid cell wall were obstacles to exploring the cell biology of this model eukaryot ...