Model organisms are of great importance to our understanding of basic biology and to making advances in biomedical research. However, the influence of laboratory cultivation on these organisms is underappreciated, and especially how that environment can affect research outcomes. Recent experiments led to insights into how the widely used laboratory reference strain of the nematode Caenorhabditis elegans compares with natural strains. Here we describe potential selective pressures that led to the fixation of laboratory-derived alleles for the genes npr-1, glb-5, and nath-10. These alleles influence a large number of traits, resulting in behaviors that affect experimental interpretations. Furthermore, strong phenotypic effects caused by these laboratory-derived alleles hinder the discovery of natural alleles. We highlight strategies to reduce the influence of laboratory-derived alleles and to harness the full power of C. elegans.

Caldicott IM et al. (1994) "Laboratory cultivation of Caenorhabditis elegans and other free-living nematodes."

Riddle DL, Caldicott IM, Larsen PL

[

1994]

Nematodes have been cultured continuously in the laboratory since 1944 when Margaret Briggs Gochnauer isolated and cultured the free-living hermaphroditic species Caenorhabditis briggsae. Work with C. briggsae and other rhabditid nematodes, C. elegans, Rhabditis anomala, and R. pellio, demonstrated the relative ease with which they could be cultured. The culturing techniques described here were developed for C. elegans, but are generally suitable (to varying degrees) for other free-living nematodes. Whereas much of the early work involved axenic culturing, most of these techniques are no longer in common use and are not included here. In the 1970s C. elegans became the predominant research model due to work by Brenner and co-workers on the genetics and development of this species. An adult C. elegans is about 1.5 mm long, and under optimal laboratory conditions has a life cycle of approximately 3 days. There are two sexes, males and self-fertile hermaphrodites, that are readily distinguishable as adults. The animals are transparent throughout the life cycle, permitting observation of cell divisions in living animals using differential interference microscopy. The complete cell lineage and neural circuitry have been determined and a large collection of behavioral and anatomical mutants have been isolated. C. elegans has six developmental stages: egg, four larval stages (L1-L4), and adult. Under starvation conditions or specific manipulations of the culture conditions a developmentally arrested dispersal stage, the dauer larva, can be formed as an alternative third larval stage. Many of the protocols included here and other experimental protocols have been summarized in "The Nematode Caenorhabditis elegans". We also include a previously unpublished method for long-term chemostat cultures of C. elegans. General laboratory culture conditions for nematode parasites of animals have been described, but none of these nematodes can be cultured in the laboratory through more than one life cycle. Marine nematodes and some plant parasites have been cultured xenically or with fungi. Laboratory cultivation of several plant parasites on Arabidopsis thaliana seedlings in agar petri plates has also been reported.

Leilani M Miller (2001) International C. elegans Meeting "USING C. ELEGANS IN THE UNDERGRADUATE LABORATORY"

Leilani M Miller

[

International C. elegans Meeting,

2001]

I have used C. elegans in two different undergraduate lab courses at Santa Clara University. One is an upper division research-focused "Project Lab" course described below and the other is an upper division Genetics course that uses C. elegans for its laboratory component. The "Project Lab" course was developed with an NSF grant to integrate research and education by designing and implementing an intensive new laboratory course in which undergraduate students participate in supervised research projects. These research projects are directly related to ongoing research in my laboratory. Project Lab has a class size of 7-12 students and, each year, it focuses on research activities appropriate for that particular stage of the research project. The addition of this course to our biology curriculum enhances the science education of undergraduate students at Santa Clara University by providing hands-on laboratory experience and exposure to real research problems. It also enhances the research program of the principal investigator by training and motivating students for research. Specifically, we are using sequencing and site-directed mutagenesis to conduct a structure/function analysis of LIN-31, a member of the winged-helix family of transcription factors, which is required for the proper specification of vulval cell fates in C. elegans . The first two Project Lab classes made considerable progress, resulting in a paper on which three Project Lab students were authors (Miller et al. , Genetics 156: 1595, 2000). Results from the two most recent Project Lab class will be presented as a poster at this meeting (see Mora-Blanco et al. ). I have also successfully used C. elegans in my upper division genetics laboratory course. This is a 10-week course in which students use C. elegans to study basic genetic principles such as dominance, independent assortment, recombination, and complementation. In order to do this, they first find and then analyze their own genetic mutant. As the quarter progresses, they use their mutants to study the basic genetic principles listed above. The first lab introduces them to C. elegans and teaches them how to grow, manipulate, and sex the animals. In the second lab, they "find" their mutant by doing a mock mutant screen. In subsequent labs, they carry out genetic manipulations (crosses) to better understand their mutants. By the end of the quarter, they can answer the following questions about their mutant: Is the mutant allele dominant or recessive to the wild-type allele? What chromosome does it map to? Between what other genes does it map? Is it an allele of a known gene? In summary, C. elegans is ideally suited for an undergraduate laboratory course. With appropriate training, undergraduate students can quickly learn to manipulate nematodes for use in genetic and/or molecular experiments. Furthermore, the flexibility of the 3-, 4-, or 7-day life cycle makes C. elegans particularly appealing for institutions on the quarter system. Access to teaching materials for both courses will be available at the teaching poster session and on my web page (www-acc.scu.edu/~lmiller/homepage.html).

Duerr, Janet S. (2013) International Worm Meeting "C. elegans modules for multiple laboratory classes."

Duerr, Janet S.

[

International Worm Meeting,

2013]

At large public universities, it is often difficult to provide students with the optimum breadth and depth of laboratory experiences. To cope with the realities of team teaching, I have developed C. elegans based modules for three different lab courses: Introductory Biology (taught to 500 freshmen per semester), Genetics Lab (~50 upperclassmen per semester), and Cell Biology Lab (16 upperclassmen per semester). These modules provide a way to use C. elegans to introduce students to multiple subjects and techniques at different levels of complexity. Introductory Biology: The most basic module takes two meetings in the middle of the first term of a joint lecture-lab course; students have studied basic genetics and are studying cells. In week 1, students receive plates of N2, dpy-5, and unc-52. The students observe and record body bends per minute using a dissecting scope and learn about mutant phenotypes. In week 2, they analyze their data using Excel and use a shared fluorescent microscope to observe and photograph GFP+ worms. Genetics Lab: The module in this team-taught lab (which also uses Drosophila, bacteria, and yeast) introduces students to RNAi and gives them more experience with fluorescent microscopy. In week 1, students receive a regular or RNAi-hypersensitive GFP+ strain and plates with OP50 or dpy-5 RNAi bacteria. In week 2, the students use upright (fluorescent) microscopes to observe the morphology of their treated worms vs. dpy-5 mutants. The students take photographs and quantify worm length using ImageJ and Excel. Cell Biology Lab: "My" lab makes extensive uses of transgenic nematodes. Each student receives a different GFP-translational fusion strain near the beginning of the course. As an introduction to bioinformatics, the students use WormBase to gather information on their strains and GFP labeled proteins. The students use their strains to run protein gels and Westerns. They use fluorescent teaching scopes with digital cameras to observe and photograph their strains. They perform indirect immunofluorescence with an anti-GFP antibody, a second antibody, and phalloidin. The students are given supervised time at the confocal microscope to take images of their strain. The final class presentations allow the students to discuss the cell biology of 'their' GFP-proteins with each other.

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