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[
Methods Cell Biol,
1995]
DNA transformation assays in a whole organism provide experimental links between molecular structure and phenotype. Experiments with transgenic Caenorhabditis elegans start in general with the injection of DNA into the adult gonad. Effects on phenotype or gene expression patterns can be analyzed either in F1 progeny derived from the injected animals or in derived transgenic lines. Microinjection of C. elegans was first carried out by Kimble et al. (1982). Stinchcomb et al. (1985) then showed that injected DNA could be maintained for several generations in transgenic lines. The first selective methods for producing and maintaining transgenic lines were reported in 1986 (Fire, 1986). These methods have been considerably improved since then (Mello et al., 1991) , so that assays involving DNA transformation are now a standard part of the experimental repertoire for C. elegans.
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[
2006]
RNA interference (RNAi) describes a conserved biological response to double-stranded RNA (dsRNA) resulting in the degradation of homologous messenger RNA. In the last few years, this process of sequence-specific, post-transcriptional gene silencing has become a key technique for rapidly assessing gene function in species ranging from plants to mammals. Fire et al. provided the first insight into the RNAi mechanism by identifying dsRNA as the trigger of RNAi in Caenorhabditis elegans in 1998 [1]. However, a similar gene-silencing phenomenon was reported in earlier studies in both plants and Neurospora [2,3]. The basic RNAi response starts with long dsRNA being processed into small interfering RNAs (siRNAs) by a ribonuclease (RNase) III enzyme, Dicer. Next, the siRNA is incorporated into the RNA-induced silencing complex (RISC). For target RNA recognition to occur, the siRNA duplex must be unwound, allowing binding of one siRNA strand to the target mRNA. This is followed by RISC cleavage of the homologous mRNA. Recent work has shown that the RNAi machinery is also involved in antiviral responses, transposon silencing, development and heterochromatin formation [4].
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[
2005]
RNA interference (RNAi) is a recently discovered phenomenon in which doublestranded RNA (dsRNA) silences endogenous gene expression in a sequencespecific manner (Fire et al., 1998). Since its discovery, the use of RNAi has become widely employed in many organisms to specifically knock down gene function. RNAi shares a remarkable degree of similarity with silencing phenomena in other organisms (Cogoni et al., 1999a; Sharp, 1999). For instance, RNAi, posttranscriptional gene silencing in plants and cosuppression in fungi can all be activated by the presence of aberrant RNAs (Maine, 2000; Tijsterman et al., 2002a). Additionally, plant, worm, and fly cells or extracts undergoing RNA-mediated interference all contain small dsRNAs, around 25 nucleotides in length, identical to the sequences present in the silenced gene (Baulcombe, 1996; Hammond et al., 2000; Zamore et al., 2000; Catalanotto et al., 2000). The high degree of similarity between these RNA-mediated silencing phenomena supports the notion that they were derived from an ancient and conserved pathway used to regulate gene expression, presumably to eliminate defective RNAs and to defend against viral infections and transposons. (Zamore, 2002). Components of RNAi have also been implicated in developmental processes, suggesting that RNAi may play a broader role in regulating gene expression (Smardon et al., 2000; Knight et al., 2001; et al., Ketting et al., 2001). Although we have learned much about the general mechanisms underlying RNAi, a detailed understanding of how RNAi works remains to be elucidated. In this chapter we will discuss first the biology of RNAi, then the genes required for its function, and we will end with a discussion on recent findings that have implicated chromatin silencing in the mechanism of RNAi.
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[
1960]
For the purpose of the present chapter the noun 'cultivation' is to be taken as the maintenance, in the laboratory, of a population of organisms belonging to a desired species through successive generations and subcultures over a prolonged period of time (weeks, months, or years). This is a deliberate restriction of the term. The noun 'culture' is most aptly used for a population within a circumscribed vessel or container (test-tube, Petri dish, U.S. Bureau of Plant Industry watch glass, etc.); it is also used in a looser, more general way (as "in culture") to cover conditions of substantial growth whether or not leading to cultivation in the strict sense
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[
Methods Cell Biol,
1995]
Geneticists like to point out that the ultimate test of a proposed function for a gene and its encoded product (or products) in a living organism involves making a mutant and analyzing its phenotype. This is the goal of reverse genetics: a gene is cloned and sequenced, its transcripts and protein coding sequence are analyzed, and a function may be proposed; one must then introduce a mutation in the gene in a living organism to see what the functional consequences are. The analysis of genetic mosaics takes this philosophy a step further. In mosaics, some cells of an individual are genotypically mutant and other cells are genotypically wild type. One then asks what the phenotypic consequences are for the living organism. This is not the same as asking what cells transcribe the gene or in what cells the protein product of the gene is to be found, but rather it is asking in what cells the wild-type gene is needed for a given function...
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[
1990]
Induction of the C. elegans vulva is a simple example of pattern formation in which the combined action of two intercellular signals specifies three cell types in a precise spatial pattern. These two signals, a graded inductive signal and a short-range lateral signal, are each mediated by a distinct genetic pathway. To understand how these intercellular signals specify cell type, we are studying, by genetic analysis and molecular cloning, genes whose products are involved in the induction pathway.
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[
2000]
Computer tracking of Caenorhabditis elegans, a free-living soil nematode, is a promising tool to assess behavioral changes upon exposure to contaminants. A short life cycle, a known genetic make-up, thoroughly studied behavior, and a completely mapped nervous system make C. elegans an attractive soil test organism with many advantages over the commonly used earthworm. Although many toxicity tests have been performed with C. elegans, the majority focused on mortality, a much less sensitive endpoint than behavior. A computer tracking system has been developed to monitor behavioral changes using C. elegans. Because conditions unrelated to specific toxicant exposures, such as changes in temperature, developmental stage, and presence of adequate food sources, can affect behavior, there is a need to standardize tracking procedures. To this end, we have developed reference charts for control movement comparing the movement of four and five day-old adult nematodes. The use of K-medium versus deionized (DI) H2O for pre-tracking rinses was also investigated. A final reference chart compared the behavioral responses of nematodes at various food densities (i.e. bacterial concentrations).
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[
Methods Cell Biol,
1995]
The number of easily distinguishable mutant phenotypes in Caenorhabditis elegans is relatively small, and this constrains the number of factors that can be followed in standard genetic crosses. Consequently, a new mutation is mapped, first to a chromosome using two-factor data from one or more crosses, and then to a chromosomal subregion by successive three-factor crosses. Mapping would be more efficient if it were possible to score a large number of well-distributed markers in a single cross. The advent of the polymerase chain reaction makes this approach feasible by allowing polymorphic genomic regions to serve as genetic markers that are easily scored in DNA released from individual animals. The only "phenotype" is a band on a gel, so the segregation of many of these markers can be followed in a single cross. Following the terminology proposed by Olsen et al. (1989), we refer to polymorphisms that can be scored by appropriately designed polymerase chain reaction (PCR) assays as polymorphic seqeunce-tagged sites (STSs)...
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[
1985]
At first sight the inclusion of a chapter on Caenorhabditis elegans in a volume on cell biology may seem unusual. However this nematode has been a superb model system for a number of cell biology studies as well as a useful model of aging. This widespread interest in C. elegans is engendered in large part by its genetic system and its optical clarity in Nomarski phase-contrast optics. Nematodes have long been a system in wide use among experimental gerontologists, and with the introduction of C. elegans by Brenner in 1974, this species has become the nematode of choice for most aging studies. We concentrate primarily on C. elegans in this review although a number of other speices, including Caenorhabditis briggsae, Turbatrix aceti, and Panagrellus redivivus, have been used in aging studies also. Other reviews on aging in C. elegans have appeared recently, including a more detailed review in another volume of this series.
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[
Lecture Notes in Computer Science,
2008]
One of the most tractable organisms for the study of nervous systems is the nematode Caenorhabditis elegans, whose locomotion in particular has been the subject of a number of models. In this paper we present a first integrated neuro-mechanical model of forward locomotion. We find that a previous neural model is robust to the addition of a body with mechanical properties, and that the integrated model produces oscillations with a more realistic frequency and waveform than the neural model alone. We conclude that the body and environment are likely to be important components of the worms locomotion subsystem.