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[
2004]
Discovered over a century ago, the centrosome is the major microtubule organizing center of the animal cell. It is a tiny organelle of surprising structural complexity. Over the last few years our understanding of the structure and composition of centrosomes has greatly advanced, and the demonstration of frequent centrosome anomalies in most common human tumors has sparked additional interest in the role of this organelle in a broader scientific community. The centrosome controls the number and distribution of microtubules.a major element of the cell cytoskeleton.and hence influences many important cellular functions and properties. These include cell shape, polarity, and motility, as well as the intracellular transport and positioning of various organelles. Of particular interest, centrosome function is critical for chromosome segregation and cell division. This book is meant to summarize our current knowledge of the structure, function and evolution of microtubule organizing centers, primarily centrosomes. Emphasis is on the role of these organelles in development and disease (particularly cancer).
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[
Methods Cell Biol,
1995]
Caenorhabditis elegans is in all likelihood the first metazoan animal whose entire genome will be determined. In addition, a very detailed description of the animal's morphology, development, and physiology is available (see elsewhere in this book, and Wood, 1988). Thus, the complete phenotype and genotype of an animal will be known. What is not known is how genotype determines phenotype; to study this, one needs to establish connections between genome sequence and phenotypes. Much has been done by classic or forward genetics: mutagenesis experiments have identified loci involved in a specific trait. Many of these loci have already been defined at the molecular level, and the genome sequence will certainly aid in the identification of many more. The opposite approach, reverse genetics, becomes naturally more important when more of the genome sequence is determined: Given the sequence of a gene of which nothing else is know, how can the function of that gene be determined? Reverse genetics is more than targeted inactivation. One can study a gene's function by several approaches...|
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[
Methods Mol Biol,
2011]
Quantitative proteomics aims to identify and quantify proteins in cells or organisms that have been obtained from different biological origin (e.g., "healthy vs. diseased"), that have received different treatments, or that have different genetic backgrounds. Protein expression levels can be quantified by labeling proteins with stable isotopes, followed by mass spectrometric analysis. Stable isotopes can be introduced in vitro by reacting proteins or peptides with isotope-coded reagents (e.g., iTRAQ, reductive methylation). A preferred way, however, is the metabolic incorporation of heavy isotopes into cells or organisms by providing the label, in the form of amino acids (such as in SILAC) or salts, in the growth media. The advantage of in vivo labeling is that it does not suffer from side reactions or incomplete labeling that might occur in chemical derivatization. In addition, metabolic labeling occurs at the earliest possible moment in the sample preparation process, thereby minimizing the error in quantitation. Labeling with the heavy stable isotope of nitrogen (i.e., (15)N) provides an efficient way for accurate protein quantitation. Where the application of SILAC is mostly restricted to cell culture, (15)N labeling can be used for micro-organisms as well as a number of higher (multicellular) organisms. The most prominent examples of the latter are Caenorhabditis elegans and Drosophila (fruit fly), two important model organisms for a range of regulatory processes underlying developmental biology. Here we describe in detail the labeling with (15)N atoms, with a particular focus on fruit flies and C. elegans. We also describe methods for the identification and quantitation of (15)N-labeled proteins by mass spectrometry and bioinformatic analysis.
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[
1990]
The free-living nematode Caenorhabditis elegans is a small and unpretentious organism. It may thrive unnoticed in the cabbage patch in your backyard or the flower pot on your balcony. In their natural habitat soil nematodes live in a thin film of water. In the laboratory C. elegans dwells on Petri dishes in the liquid film on the top of an agar layer, but can also be grown in liquid culture. As in other nematodes the liquid-filled body cavity (pseudocoelom) functions as a hydroskeleton. When the worm dries out, the hydroskeleton collapses and the animal inevitably dies. In a loose sense C. elegans may therefore be considered as a kind of aquatic animal. Because of this and because C. elegans is particularly well suited to the study of certain aspects of development, the following chapter is included in this book on Experimental Embryology of Aquatic Organisms. The intention of this contribution is to serve as an introduction and as a reference source rather than as a complete summary of present knowledge in the field. As indicated by the title, the review will focus on embryonic cell lineages, pattern formation in the embryo and the analysis of mutants affecting early
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[
2003]
Since the publication of the first genetic research paper on Caenorhabditis elegans (C. elegans for short) in 1974, this microscopic, free-living nematode has become a popular model organism to study development, neurobiology, and other biological problems. The ability to do powerful genetics has been the most critical reason why studies using this organism have made enormous contributions to basic biology and medical science. Therefore, C. elegans genetics should be part of any modern genetic education. In this chapter, we describe some of the unique properties of C. elegans that makes it an exceptional organism for genetic and molecular biological research. Some important genetic tools and methodologies developed by C. elegans researchers will also be introduced. We aim to connect the fundamental principles of genetics as described in early chapters with practical applications of these principles in actual research. We have chosen a few genetic pathways and biological problems as examples for illustrating the logic behind the genetic analyses and for introducing some commonly practiced strategies and methods. We do not hesitate to introduce some of the most advanced and up-to-date methods and approaches, including those developed since the genome sequence was determined in 1998. We believe today's students can go right into the heart of present research after learning the basic principle of Genetics (see the early chapters of this book) and molecular biology. In fact, in many C. elegans laboratories, undergraduate students are doing a wide variety of experiments using the genetic techniques