Today’s Goals Describe the advantages of C. elegans as a model organism Discuss the life cycle of the nematode Safely and effectively culture a population of C. elegans and transfer worms to new plates for experimental studies

Caenorhabditis elegans The model organism: Caenorhabditis elegans Electron micrograph of a C. elegans hermaphrodite

Can be male or hermaphroditic Small = about 1mm in size Lives in the soil Nematode Eats bacteria Can be male or hermaphroditic Self-fertilizing!

Caenorhabditis elegans Profile Soil nematode, Eukaryote Genome size: 100 Mb Number of chromosomes: 6 Generation time: about 2 days Female reproductive capacity: 250 to 1000 progeny Special characteristics Strains Can Be Frozen Easy to grow Hermaphrodite Known cell lineage pattern for all 959 somatic cells Only 302 neurons Transparent body Can be characterized genetically About 70% of Human Genes have related genes in C. elegans

Lifecycle of a worm (from www.wormatlas.org)

Anatomy of a worm (from www.wormatlas.org)

C. elegans cell division can be studied in the transparent egg

C. elegans cell lineage is known

C. elegans is amenable to many forms of RNAi treament Fire & Mello - Won Nobel prize for discovery of RNAi Feeding worms bacteria that express dsRNAs or soaking worms in dsRNA sufficient to induce silencing (Gene 263:103, 2001; Science 282:430, 1998)

C. elegans can also be used for Behavioral studies Will respond when tapped - Different mutants might respond differently Display chemotaxis Move toward certain chemicals Studying Neurobiology Very few neurons Easy to map out what neurons target each part of the body, test genetics of neurobiology Studying Embryonic Development Cell biology, Apoptosis . . . And more !!! C. Elegans in space!!!

Nobel Prize winners who did their research with C. elegans Sydney Brenner John Sulston Bob Horvitz Andrew Fire Craig Mello

http://park. itc. utokyo. ac. jp/mgrl/IINO_lab/Eng_figures/Fig4 http://park.itc.utokyo.ac.jp/mgrl/IINO_lab/Eng_figures/Fig4.JPEG&imgrefurl=http://park.itc.utokyo.ac.jp/mgrl/IINO_lab/IINO_lab.html&h=491&w=333&sz=29&hl=en&start=23&um=1&tbnid=6D_KWwsfwR_pZM:&tbnh=130&tbnw=88&prev=/images%3Fq%3Dc.%2Belegans%2Bchemotaxis%26start%3D20%26ndsp%3D20%26um%3D1%26hl%3Den%26client%3Dsafari%26rls%3Den%26sa%3DN We have found a novel form of behavioral plasticity in chemotaxis. That is, worms starved in the presence of NaCl learn to avoid NaCl, which is normally an attractant for C. elegans. Because starvation in the absence of NaCl has little or no effect on chemotaxis, this seems to be an associative learning. While similar bahavioral changes were observed using several other water-soluble chemoattractants, chemotaxis to odorants is not affected. We have isolated several mutants defective in this form of learning and also identified several mutants already known for other defects to have defects in learning in our paradigm. We plan to clone or analyze the genes responsible for these defects.

How will we be working with C. elegans? We will be following along with lab protocols from www.silencinggenomes.org Learn to culture and examine nematodes Learn techniques to count and transfer worms to separate dishes Perform RNAi, using a kit Design experiments of our own with techniques we read about in the primary literature

We can order C. elegans Mutants University of Minnesota Caenorhabditis genetics center 7$ per strain http://www.cbs.umn.edu/CGC/

NGM Plates and OP50 broth You will pour NGM-lite plates Pour 1 bottle per pair of students Using Sterile Technique Nematode Growth Medium Label Plates NGM, Date, Initials When solid, invert, put on lab bench by incubators Then – take 5 microliters of OP50 bacteria, innoculate a tube of LB broth (on lab bench) Label with your initials, and OP50, put in 37 Degree incubator Next time - we’ll seed plates with bacteria for culture of C. elegans