-
[
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.
-
[
WormBook,
2005]
Genetic suppression has provided a very powerful tool for analyzing C. elegans. Suppression experiments are facilitated by the ability to handle very large numbers of individuals and to apply powerful selections. Because the animal grows as a self-fertilizing diploid, both dominant and recessive suppressors can be recovered. Many different kinds of suppression have been reported. These are discussed by category, with examples, together with discussion of how suppressors can be used to interpret the underlying biology, and to enable further experimentation. Suppression phenomena can be divided into intragenic and extragenic classes, depending on whether the suppressor lies in the same gene as the starting mutation, or in a different gene. Intragenic types include same-site replacement, compensatory mutation, alteration in splicing, and reversion of dominant mutations by cis- knockout. Extragenic suppression can occur by a variety of informational mechanisms, such as alterations in splicing, translation or nonsense-mediated decay. In addition, extragenic suppression can occur by bypass, dosage effects, product interaction, or removal of toxic products. Within signaling pathways, suppression can occur by modulating the strength of signal transmission, or by epistatic interactions that can reveal the underlying regulatory hierarchies. In C. elegans biology, the processes of muscle development, vulva formation and sex determination have provided remarkably rich arenas for the investigation and exploitation of suppression.
-
[
Genetics,
2022]
Over the last 20 years, studies of Caenorhabditis elegans natural diversity have demonstrated the power of quantitative genetic approaches to reveal the evolutionary, ecological, and genetic factors that shape traits. These studies complement the use of the laboratory-adapted strain N2 and enable additional discoveries not possible using only one genetic background. In this chapter, we describe how to perform quantitative genetic studies in Caenorhabditis, with an emphasis on C. elegans. These approaches use correlations between genotype and phenotype across populations of genetically diverse individuals to discover the genetic causes of phenotypic variation. We present methods that use linkage, near-isogenic lines, association, and bulk-segregant mapping, and we describe the advantages and disadvantages of each approach. The power of C. elegans quantitative genetic mapping is best shown in the ability to connect phenotypic differences to specific genes and variants. We will present methods to narrow genomic regions to candidate genes and then tests to identify the gene or variant involved in a quantitative trait. The same features that make C. elegans a preeminent experimental model animal contribute to its exceptional value as a tool to understand natural phenotypic variation.
-
[
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.
-
[
Genetics,
2018]
Since the earliest days of research on nematodes, scientists have noted the developmental and morphological variation that exists within and between species. As various cellular and developmental processes were revealed through intense focus on <i>Caenorhabditis elegans</i>, these comparative studies have expanded. Within the genus <i>Caenorhabditis</i>, they include characterization of intraspecific polymorphisms and comparisons of distinct species, all generally amenable to the same laboratory culture methods and supported by robust genomic and experimental tools. The <i>C. elegans</i> paradigm has also motivated studies with more distantly related nematodes and animals. Combined with improved phylogenies, this work has led to important insights about the evolution of nematode development. First, while many aspects of <i>C. elegans</i> development are representative of <i>Caenorhabditis</i>, and of terrestrial nematodes more generally, others vary in ways both obvious and cryptic. Second, the system has revealed several clear examples of developmental flexibility in achieving a particular trait. This includes developmental system drift, in which the developmental control of homologous traits has diverged in different lineages, and cases of convergent evolution. Overall, the wealth of information and experimental techniques developed in <i>C. elegans</i> is being leveraged to make nematodes a powerful system for evolutionary cellular and developmental biology.
-
[
WormBook,
2006]
Through genetic analyses, the function of genes is investigated by studying organisms where gene function is altered. In classical forward genetic screening, individuals are treated with mutagens to induce DNA lesions and mutants with a phenotype of interest are sought. After a mutant is found, the gene mutated is identified through standard molecular techniques. Detailed studies of the mutant phenotype coupled with molecular analyses of the gene allows elucidation of the gene's function. Forward genetics has been responsible for our understanding of many biological processes and is an excellent method for identifying genes that function in a particular process.In reverse genetics, the functional study of a gene starts with the gene sequence rather than a mutant phenotype. Using various techniques, a gene's function is altered and the effect on the development or behaviour of the organism is analysed. Reverse genetics is an important complement to forward genetics. For example, using reverse genetics, one can investigate the function of all genes in a gene family, something not easily done with forward genetics. Further, one can study the function of a gene found to be involved in a process of interest in another organism, but for which no forward genetic mutants have yet been identified. Finally, the vast majority of genes have not yet been mutated in most organisms and reverse genetics allows their study. The availability of complete genome sequences combined with reverse genetics can allow every gene to be studied.This chapter gives detailed protocols for the two main methods of perturbing gene function in C. elegans: RNA interference and the creation of deletion mutants. Either technique can be applied to the study of individual genes. With less than a day of actual work, RNAi creates a knockdown of gene function without altering the organism's DNA (see below). In contrast, with about a month of work, a deletion mutation permanently removes all gene function. Deciding which technique to use will depend on the nature of the experiment. The techniques can also be combined, where RNAi is used for rapid screening of loss of function phenotypes and then deletion mutants are made to study genes of particular interest. RNAi can also be carried out on a global scale, where knockdown of (nearly) every gene is tested for inducing a phenotype of interest. In this case, the reverse genetics technique of RNAi can be thought of as a forward genetic screening tool.
-
[
WormBook,
2005]
Evolutionary innovation requires genetic raw materials upon which selection can act. The duplication of genes is of fundamental importance in providing such raw materials. Gene duplications are very widespread in C. elegans and appear to arise more frequently than in either Drosophila or yeast. It has been proposed that the rate of duplication of a gene is of the same order of magnitude as the rate of mutation per nucleotide site, emphasising the enormous potential that gene duplication has for generating substrates for evolutionary change. The fate of duplicated genes is discussed. Complete functional redundancy seems unstable in the long term. Most models require that equality amongst duplicated genes must be disrupted if they are to be preserved. There are various ways of achieving inequality, involving either the nonfunctionalization of one copy, or one copy acquiring some novel, beneficial function, or both copies becoming partially compromised so that both copies are required to provide the overall function that was previously provided by the single ancestral gene. Examples of C. elegans gene duplications that appear to have followed each of these pathways are considered.
-
[
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.
-
[
WormBook,
2007]
Sexual identity is one of a few basic parameters that specify how development should proceed. Although sex determination has profound effects on many tissues, its most ancient and fundamental role is ensuring that some germ cells become sperm, and others become oocytes or eggs. Spermatocytes and oocytes are usually produced in male and female animals, respectively, but C. elegans is uniquely suitable for studying the control of these cell fates because both types of cells are made from a common pool of progenitors in XX hermaphrodites. Extensive genetic and molecular studies have shown that the sexual fate of germ cells in C. elegans is controlled by the same genes that regulate sexual identity in other parts of the animal. However, this regulatory pathway has additional features that are unique to the germ line. First, several genes, like the three fogs, act only in germ cells. Second, the three fem genes act in concert with targets of
tra-1 to control germ cell fates, but do not act this way in the soma. Third, translational repression of
tra-2 is essential for hermaphrodite spermatogenesis. Fourth, translational repression of
fem-3 is needed for oogenesis. In this review, we present genetic and molecular models for how these processes work, and summarize the evidence upon which they are built.
-
[
WormBook,
2007]
Strongyloides is a genus of parasitic nematodes, which, unusually, has a free-living adult generation. Here we introduce the biology of this genus, especially the fascinating, but complex, life-cycle together with an overview of the taxonomy, morphology, genetics and genomics of this genus.