Laboratory cultivation of caenorhabditis elegans and other free-living nematodes

This chapter describes the cultivation of caenorhabditis elegans and other free-living nematodes in the laboratory. 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 the observation of cell divisions in living animals using a differential interference microscopy. Tools for manipulating individual nematodes on plates are made by anchoring a 1.5-cm piece of 32-gauge platinum wire in a 6 inch inoculating loop holder or in a Pasteur pipette. For best results the end of the wire used for manipulation should be flattened and, if necessary, rounded to remove sharp edges. Remove supernatant and re-suspend in 15 ml of M9 buffer, divide between two 15-ml tubes, and place on ice. When cold, add 7.5 ml of cold 60% sucrose to each tube. Mix by inversion and centrifuge immediately at 1500 RCF for 5 min.