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[
Methods Cell Biol,
2013]
Lipid droplets (LDs) are an intracellular organelle, consisting of a neutral lipid core covered by a monolayer of phospholipids and proteins. It primarily mediates lipid storage, metabolism, and transportation. Recently, research of LDs has emerged as a rapidly developing field due to the strong linkage between ectopic lipid accumulation and metabolic syndromes. Recently, more than 30 proteomic studies of isolated LDs have identified many important LD proteins that have highlighted and have also predicted the potential biological roles of the organelle, motivating the field to develop quite rapidly. This chapter summarizes methods used in proteomic studies for three representative species reported and discusses their advantages and disadvantages. We believe that this chapter provides useful information and methods for future LD proteomic studies especially for LDs in other species.
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[
Modeling in Molecular Biology. G Ciobanu and G Rozenberg (eds). Natural Computing Series, Springer-Verlag.,
2004]
We present preliminary results of a new approach to the formal modeling of biological phenomena. The approach stems from the conceptual compatibility of the methods and logic of data collection and analysis in the field of developmental genetics with the languages, methods and tools of scenario-based reactive system design. In particular, we use the recently developed methodology consisting of the language of live sequence charts with the play-in/play-out process, to model the well-characterized process of the cell fate acquisition during C. elegans
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[
WormBook,
2005]
Synaptogenesis is a process involving the formation of a neurotransmitter release site in the presynaptic neuron and a receptive field at the postsynaptic partners, and the precise alignment of pre- and post-synaptic specializations. In C. elegans synapses are found as en passant axonal swellings along the nerve processes. Genetic screens using a synaptic vesicle-associated GFP marker have identified key players in synaptic target recognition and organization of the presynaptic terminals. Importantly, the functions of most genes are evolutionarily conserved. Further studies using a combination of genetic modifier screens and reverse genetics have begun to reveal the underlying signaling pathways.
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[
2013]
One of the central aims of modern biology is to elucidate the mechanisms leading to the complex structures of organisms. Using the information inherent in the one-dimensional nucleotide sequence of the genome, cleavage divisions of the fertilized egg, with all the associated genetic and epigenetic regulatory steps, generates a specific three-dimensional pattern of differentiated cells. Anensuing series of dynamic processes finally results in a functional organism with many complex structures and phenotypes. Understanding the genetic and molecularbasis of embryonic and postembryonic patterning during development has been a central field of research since the 1970s, and nematodes have been at the forefront of this research.
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[
Methods Cell Biol,
2012]
In Caenorhabdatis elegans as in other animals, fat regulation reflects the outcome of behavioral, physiological, and metabolic processes. The amenability of C. elegans to experimentation has led to utilization of this organism for elucidating the complex homeostatic mechanisms that underlie energy balance in intact organisms. The optical advantages of C. elegans further offer the possibility of studying cell biological mechanisms of fat uptake, transport, storage, and utilization, perhaps in real time. Here, we discuss the rationale as well as advantages and potential pitfalls of methods used thus far to study metabolism and fat regulation, specifically triglyceride metabolism, in C. elegans. We provide detailed methods for visualization of fat depots in fixed animals using histochemical stains and in live animals by vital dyes. Protocols are provided and discussed for chloroform-based extraction of total lipids from C. elegans homogenates used to assess total triglyceride or phospholipid content by methods such as thin-layer chromatography or used to obtain fatty acid profiles by methods such as gas chromatography/mass spectrometry. Additionally, protocols are provided for the determination of rates of intestinal fatty acid uptake and fatty acid breakdown by -oxidation. Finally, we discuss methods for determining rates of de novo fat synthesis and Raman scattering approaches that have recently been employed to investigate C. elegans lipids without reliance on invasive techniques. As the C. elegans fat field is relatively new, we anticipate that the indicated methods will likely be improved upon and expanded as additional researchers enter this field.
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[
1983]
In the preparation of this review, I have made the basic assumption that the desire of the reader is to understand the biological basis of organismic aging. Given this premise, the organism of choice should be one that offers the most immediate hope of arriving at such an understanding. An ideal organism should have a short lifespan; be inexpensive to maintain; be experimentally malleable by a variety of techniques including molecular, morphological, genetic, and biological approaches; and be the object of study in a sufficient number of different laboratories to assure the accumulation of a critical mass of data. The nematode, Caenorhabditis elegans, admirably fulfills all of these basic requirememts. Researchers in the field of aging are faced with a large number of different theories which purport to explain the molecular basis of organismic aging. There are two major reasons for this proliferation of theoretical views. First, aging is an extremely complex phenomenon involving changes in a number of different physiological systems; these physiological changes are often detected, but proof that any one of the changes is responsible for aging is lacking. Second, the focus of a great deal of the research in the field has not been so much on understanding the biological basis of the entire aging process as on understanding one or another of the consequences of this process, particularly in humans and other mammals. The mammalian model systems may often be quite inappropriate for addressing the more basic, long-term questions about the
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Embryos are unique in their combination of pluripotency, three-dimensionality, and the swiftness of subcellular and developmental rearrangements. In some species, it is possible to observe the entire organism taking form within a microscope field. Capturing the spatial and temporal dynamic complexity of embryogenesis tests the limits of both culture and microscopy techniques. Observing specific fluorescently labeled components during embryonic development promises to reveal the roles of organelles and molecules in a native and reproducible context. However, to gain a thorough understanding of such dynamic biological systems, one must record events of interest as they occur, while limiting the perturbations caused by the observation techniques. In this chapter, we discuss our experiences using the relatively new technology of multiphoton laser scanning microscopy to examine the development of mammalian and nematode embryos in four dimensions.
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[
1975]
Studies in behaviour genetics have covered a wide field: motivation, development, sensory capacities, intelligence, learning, evolution, neuromorphology and neurochemistry have all been approached using genetic techniques, and there are probably others. Whilst it is at present impossible to construct any unities one must accept that many such studies have as their common aim one of the most fundamental problems in biology: how is behavioral potential encoded in genetic terms and expressed in the course of development? The relative enormity of this problem is often matched by its inaccessibilty. It cannot be claimed that there is any agreed view of the way forward and much of the work has frankly to be opportunistic-seizing on some favourable material or a useful new analytical technique to gain a limited objective. Consequently, behaviour genetics often presents a confusing picture of numerous disjointed studies, with
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[
1998]
The use of antibodies to visualize the distribution and subcellular localization of gene products powerfully complements genetic and molecular analysis of gene function in C. elegans. The challenge to immunolabeling C. elegans is finding the fixation and permeabilization methods that effectively make antigens accessible without destroying the tissue morphology or the antigen. Embryos are surrounded by a chitinous eggshell and larvae and adults are surrounded by a collagenous cuticle, each of which must be permeabilized to allow penetration of antibodies. In addition, antigens and antibodies are sensitive to different fixing and permeabilizing conditions. For example, some antibodies do not work well on paraformaldehyde-fixed samples, and others are sensitive to incubation in acetone. There are many protocols used in the C. elegans field; additional protocols are summarized in Miller and Shakes (1994) and on the C. elegans World
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[
1998]
The use of antibodies to visualize the distribution and subcellular localization of gene products powerfully complements genetic and molecular analysis of gene function in C. elegans. The challenge to immunolabeling C. elegans is finding the fixation and permeabilization methods that effectively make antigens accessible without destroying the tissue morphology or the antigen. Embryos are surrounded by a chitinous eggshell and larvae and adults are surrounded by a collagenous cuticle, each of which must be permeabilized to allow penetration of antibodies. In addition, antigens and antibodies are sensitive to different fixing and permeabilizing conditions. For example, some antibodies do not work well on paraformaldehydefixed samples, and others are sensitive to incubation in acetone. There are many protocols used in the C. elegans field; additional protocols are summarized in Miller and Shakes (1994) and on the C. elegans World Wide Web page
(http://elegans.swmed.edu/).