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[
WormBook,
2005]
Mutations in many genes can result in a similar phenotype. Finding a number of mutants with the same phenotype tells you little about how many genes you are dealing with, and how mutable those genes are until you can assign those mutations to genetic loci. The genetic assay for gene assignment is called the complementation test. The simplicity and robustness of this test makes it a fundamental genetic tool for gene assignment. However, there are occasional unexpected outcomes from this test that bear explanation. This chapter reviews the complementation test and its various outcomes, highlighting relatively rare but nonetheless interesting exceptions such as intragenic complementation and non-allelic non-complementation.
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[
Methods Cell Biol,
1995]
This chapter is devoted to providing information on techniques applicable to studying transcription and translation in Caenorhabditis elegans. These techniques are constantly evolving and being passed among workers, each making improvements or adaptations. None of the techniques discussed below are original, but, rather, have emerged from a variety of sources over the years, making it difficult to trace their origin or give credit to the originators. Although each technique has been used successfully, for each there are alternative methods available in the literature that work equally well. In fact, depending on the available resources, you might find that an alternative technique suits your needs and facilities better than the one described below. For this reason, the procedures discussed below are usually accompanied by one or more references that will allow you to look at other, related methods. Where appropriate, there will also be a discussion of factors to consider when
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[
WormBook,
2007]
Because of their free-living life cycle alternatives, Strongyloides and related nematode parasites may represent the best models for translating C. elegans science to the study of nematode parasitism. S. stercoralis, a significant pathogen of humans, can be maintained in laboratory dogs and gerbils. Biosafety precautions necessary for work with S. stercoralis, though unfamiliar to many C. elegans researchers, are straightforward and easily accomplished. Although specialized methods are necessary for large-scale culture of the free-living stages of S. stercoralis, small-scale cultures for experimental purposes may be undertaken using minor modifications of standard C. elegans methods. Similarly, the morphological similarities between C. elegans and the free-living stages of S. stercoralis allow investigational methods such as laser cell ablation and DNA transformation by gonadal microinjection to be easily adapted from C. elegans to S. stercoralis. Comparative studies employing these methods have yielded new insights into the neuronal control of the infective process in parasites and its similarity to regulation of dauer development in C. elegans. Furthermore, we have developed a practical method for transient transformation of S. stercoralis with vector constructs having various tissue- and cell-specific expression patterns and have assembled these into a modular vector kit for distribution to the community.
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[
1981]
A neuron can be characterized by its morphology, transmitter (s?), receptor(s) and the nature of its synaptic contacts (chemical or electrical; excitatory or inhibitory; number and distribution of synapses; identity of the cells to which it is presynaptic or postsynaptic). It is clear that according to such criteria nervous sytems consist of neurons of many distinct types. The origin of neuronal diversity is unknown. Both how such diversity is generated during development and how the relevant developmental programme is encoded in the genome remain to
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[
Methods Cell Biol,
1995]
Complementary DNA libraries are useful tools for uncovering genes of interest in C. elegans and finding specific homologies to genes in other organisms (Waterston et al., 1992; McCombie et al., 1992). When working with existing cDNA libraries, be sure to carefully choose which libraries would be most beneficial to the type of research being done. Some libraries may be specific for genes that are present in lower copy numbers, whereas others may be of a more general nature. It is important to fully understand the source and construction of the library you will be working with. Once an appropriate library has been chosen, work may begin to isolate a specific cDNA and sequence it completely or to survey many cDNAs by single-pass DNA sequencing. Whatever the project, it is important to develop a specific strategy for both the sequencing and the organization of the clones being characterized. The strategies and procedures we have outlined in this chapter have proven effective for rapid and comprehensive cDNA characterization.
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[
2010]
Over 30 years ago, Nobel laureate Sydney Brenner recognized that an intellectually straightforward strategy to delineate the basic principles in neurobiology is to utilize a model organism with a nervous system that is simple enough to lend itself to anatomical, cellular, genetic, and molecular analysis, yet be complex enough that lessons learned in that organism would give us insight into general principles of neural function. The humble organism he chose, the nematode Caenorhabditis elegans, is now one of the most thoroughly characterized metazoans, particularly in terms of its nervous system. One of Brenner's motivations in adapting C. elegans as a model organism was to understand the totality of the molecular and cellular basis for the control of animal behavior (Brener 1988). In this chapter, we review what is arguably the best-studied aspect of C. elegans behavior: response to chemical stimuli. The C. elegans neurobiology literature can be intimidating for the uninitiated; we attempt to limit the use of "worm jargon" in this review. For a more C. elegans-centric review, we refer you to other excellent sources (Bargmann 2006).
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[
WormBook,
2005]
Cell-division control affects many aspects of development. Caenorhabditis elegans cell-cycle genes have been identified over the past decade, including at least two distinct Cyclin-Dependent Kinases (CDKs), their cyclin partners, positive and negative regulators, and downstream targets. The balance between CDK activation and inactivation determines whether cells proceed through G 1 into S phase, and from G 2 to M, through regulatory mechanisms that are conserved in more complex eukaryotes. The challenge is to expand our understanding of the basic cell cycle into a comprehensive regulatory network that incorporates environmental factors and coordinates cell division with growth, differentiation and tissue formation during development. Results from several studies indicate a critical role for CKI-1 , a CDK inhibitor of the Cip/Kip family, in the temporal control of cell division, potentially acting downstream of heterochronic genes and dauer regulatory pathways.
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[
WormBook,
2006]
The C. elegans genome encodes many RNA-binding proteins (RBPs) with diverse functions in development, indicative of extensive layers of post-transcriptional control of RNA metabolism. A number of C. elegans RBPs have been identified by forward or reverse genetics. They tend to display tissue-specific mutant phenotypes, which underscore their functional importance. In addition, several RBPs that bind regulatory sequences in the 3'' untranslated regions of mRNAs have been identified molecularly. Most C. elegans RBPs are conserved throughout evolution, suggesting that their study in C. elegans may uncover new conserved biological functions. In this review, we primarily discuss RBPs that are associated with well-characterized mutant phenotypes in the germ line, the early embryo, or in somatic tissues. We also discuss the identification of RNA targets of RBPs, which is an important first step to understand how an RBP controls C. elegans development. It is likely that most RBPs regulate multiple RNA targets. Once multiple RNA targets are identified, specific features that distinguish target from non-target RNAs and the type(s) of RNA metabolism that each RBP controls can be determined. Furthermore, one can determine whether the RBP regulates all targets by the same mechanism or different targets by distinct mechanisms. Such studies will provide insights into how RBPs exert coordinate control of their RNA targets, thereby affecting development in a concerted fashion.
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[
1992]
In vertebrates, acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are polymorphic enzymes presenting both globular and asymmetric forms. In invertebrates, only AChE has been characterized so far that presents a reduced molecular diversity. In insects for example the major molecular form of AChE is an amphiphilic dimeric form attached to the membrane through a glycolipid covalently linked at the C-terminus of each catalytic subunit. This AChE has a substrate specificity intermediate to those of mammalina AChE and BChE. A glycoplipid-anchored 7.5S from has also been observed in the trematode Schistosoma mansoni. Asymmetric forms have never been convincingly reported in invertebrates except in the more evolved animals such as Amphioxius. In the latter case also there is no BChE but AChE presents catalytic properties intermediate to those of vertebrate AChE and BChE. We are now interested in nematode AChE(s) for the following reasons: -several species are agricultural pest and it is important to get further informations on the target of potential nematicides; -it has been shown that at least three different genes code for AChE in Caenorhabditis elegans. It is therefore interesting to see whether the presence of multiple genes results in an increased molecular diversity, to define what are the structural characteristics of each gene product and finally to clone and sequence thee three genes for evolutionary relationships with the other members of the cholinesterase
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[
WormBook,
2006]
Transposons are discrete segments of DNA capable of moving through the genome of their host via an RNA intermediate in the case of class I retrotransposon or via a "cut-and-paste" mechanism for class II DNA transposons. Since transposons take advantage of their host''s cellular machinery to proliferate in the genome and enter new hosts, transposable elements can be viewed as parasitic or "selfish DNA". However, transposons may have been beneficial for their hosts as genome evolution drivers, thus providing an example of molecular mutualism. Interactions between transposon and C. elegans research were undoubtedly mutualistic, leading to the advent of needed genomic tools to drive C. elegans research while providing insights into the transposition field. Tc1, the first C. elegans transposon to be identified, turned out to be the founding member of a widespread family of mobile elements: the Tc1/ mariner superfamily. The investigation into transposition regulation in C. elegans has uncovered an unforeseen link between transposition, genome surveillance and RNA interference. Conversely, transposons were utilized soon after their identification to inactivate and clone genes, providing some of the first molecular identities of C. elegans genes. Recent results suggest that transposons might provide a means to engineer site-directed mutations into the C. elegans genome. This article describes the different transposons present in the C. elegans genome with a specific emphasis on the ones that proved to be mobile under laboratory conditions. Mechanisms and control of transposition are discussed briefly. Some tools based on the use of transposons for C. elegans research are presented at the end of this review.