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Programmed cell death is a common cell fate in most if not all multicellular organisms. Apoptosis, which will be used as a synonym for programmed cell death throughout this chapter, occurs extensively during development as well as during later life. The development of the nematode worm Caenorhabditis elegans provides a good example of the extensive use of programmed cell death.
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[
Methods Cell Biol,
1995]
Sequence analysis of cosmids from C. elegans and other organisms currently is best done using the random or "shotgun" strategy (Wilson et al., 1994). After shearing by sonication, DNA is used to prepare M13 subclone libraries which provide good coverage and high-quality sequence data. The subclones are assembled and the data edited using software tools developed especially for C. elegans genomic sequencing. These same tools facilitate much of the subsequent work to complete both strands of the sequence and resolve any remaining ambiguities. Analysis of the finished sequence is then accomplished using several additional computer tools including Genefinder and ACeDB. Taken together, these methods and tools provide a powerful means for genome analysis in the nematode.
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[
1985]
Since the last review in this series, several important projects relating to aging research in Caenorhabditis elegans have been completed. A more detailed review of the field is available. A major focus of research in Caenorhabditis elegans over the last few years has been on development, particularly the cell lineage. The entire cell lineage of the adult hermaphrodite has been described. The genetic loci coding for myosin, for rRNA, for actin, collagen, and oocyte yolk proteins, and a major family of proteins synthesized in the sperm have been isolated using recombinant DNA techniques. A transposable element has been identified, and studies aimed at using this element as a mutagen are underway. A good start has been made in generating an ordered series of overlapping recombinant clones of the entire genome; several labs are developing techniques for transformation of the worm. Aging research has also made progress over the last few years. Single-gene mutants and selectively bred stocks displaying longer lifespans have been isolated. A number of new markers of senescence have been described. Programmed cell death during development of the worm has been a major focus of research, and mutants altering this process have been isolated. There are still a few problems for aging research: there is not a single agreed-upon method of culturing biochemical quantities of worms that also gives lifespans comparable to those of small-scale cultures; and observed differences in aging parameters that are general versus those that are due to culture conditions are still under dispute. Two methods of growth (axenic and monoxenic) are still commonly used, for the most part always in distinct laboratories. All of these findings will be described within.
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[
WormBook,
2005]
C. elegans is a member of a group of nematodes called rhabditids, which encompasses a large number of ecologically and genetically diverse species. A new, preliminary phylogenetic analysis is presented for concatenated sequences of three nuclear genes for 48 rhabditid and diplogastrid species (including 10 Caenorhabditis species), as well as four species representing the outgroup. Although many relationships are well-resolved, more data are still needed to resolve some key relationships, particularly near the base of the rhabditid tree. There is high confidence for two major clades: (1) a clade comprising Mesorhabditis Parasitorhabditis, Pelodera, Teratorhabditis plus a few other species; (2) a large clade (Eurhabditis) comprising most of the remaining rhabditid genera, including Caenorhabditis and its sistergroup Protorhabditis-Prodontorhabditis-Diploscapter. Eurhabditis also contains the parasitic strongylids, the entomopathogenic Heterorhabditis, and the monophyletic group Oscheius which includes the satellite model organism O. tipulae. The relationships within Caenorhabditis are well resolved. The analysis also suggests that rhabditids include diplogastrids, to which the second satellite model organism Pristionchus pacificus belongs. Genetic disparity within Caenorhabditis is as great as that across vertebrates, suggesting Caenorhabditis lineages are quickly evolving, ancient, or both. The phylogenetic tree can be used to reconstruct evolutionary events within rhabditids. For instance, the reproductive mode changed multiple times from gonochorism to hermaphroditism, but only once from hermaphroditism to gonochorism. Complete retraction of the male tail tip, leading to a blunt, peloderan tail, evolved at least once. Reversions to unretracted tail tips occurred within both major rhabditid groups. The phylogeny also provides a guide to species which would be good candidates for future genome projects and comparative studies.
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A previous chapter in this series (1) described, primarily, the physical mapping of the 100 Mb Caenorhabditis elegans genome by fingerprinting of cosmid clones, and the linking of the contigs thus derived by YAC hybridization. At that time, the primary function of the map was to enhance the molecular genetics of the organism by facilitating the cloning of known genes, and to serve as an archive for genomic information. However, a clonal physical map - even with good alignment to the genetic map - carries only a tiny proportion of the information present in the genome. Consequently, the current objective of the C. elegans genome project (2) is to establish of the entire genomic sequence. The bacterial clone map, although incomplete by virtue of the uncloneability of regions of the genome in cosmid vectors (a factor which we shall discuss later in this chapter), has proved a sound basis for the systematic sequence analysis. The sevenfold cosmid coverage has a resolution sufficient to enable the selection of a subset of cosmids for sequencing such that, on average, each clone contributes 30 kb of unique sequence to the whole. Sequencing projects based on bacterial clone maps (3-5) of a number of other genomes of a range of sizes are also well advanced, in particular Saccharomyces cerevisiae (15 Mb; complete), Schizosaccharomyces pombe (15Mb), and Drosohpila melanogaster (150 Mb). Although it has recently been demonstrated that small bacterial genomes can be sequenced by direct shotgun sequence analysis of the entire genome with no prior mapping (6), the ability to interrelate and map clone sets, whether derived by random selection of in a directed manner, is still the most convenient route to the sequence analysis of larger genomes.