05 - Heavy Metal: Effects of Environmental Toxins on C. elegans
Module II: Cultivation of C. elegans (“Chunking” the Plates) Excerpts from Edvo-Kit #856
In this module, you will be “chunking” your C. elegans strains. This means that you will transfer wild-type and ADR-2 mutant C. elegans to the plates you prepared in Module I. Over the next few days, both strains will establish new populations in their individual plates. Once these populations have expanded to at least 50 individuals you will continue to Module III.
REPEAT steps 2-3 for mutant strain.
Wild-type
Mutant
Seeded NGM Plates
Instructor Provided NGM plate
WAIT
INCUBATE 2-3 days.
1. LABEL the bottom of the seeded NGM plates (from Module I) with “Wild-type” or “Mutant”. 2. Your instructor will have NGM plates containing wild-type or mutant C. elegans . Using a sterile loop, CUT OUT a small portion of the NGM plate containing the wild-type strain (~1 cm square). Make sure to completely cut the agar by pushing the loop all the way to the bottom plate. 3. REMOVE the “chunk” from the plate. PLACE the chunk, worm side down, in the center of the Module I Wild-type plate. 4. With a new sterile loop, REPEAT step 2 and 3 for the mutant strain. 5. After 5-10 minutes, use a microscope to CONFIRM the presence of C. elegans on the “chun- ked” plates. NOTE: Occasionally, a “chunk” is transferred nematode side up. Worms in this position will eventually migrate to the plate. If the “chunk” is incorrectly positioned, keep the plate but wait 1 hour before confirming the presence of C. elegans and continuing on to Step 6. 6. COVER and PLACE the plates into a cardboard box. INCUBATE at room temperature for 2-3 days.
OPTIONAL STOPPING POINT: Plates may be stored for up to a week but need extra OP50 to avoid drying and to feed the growing population. See Appendix A.
7. CHECK growth of C. elegans under a microscope. If the plate contains 50 or more worms, proceed with Module III. If the plate contains fewer than 50 worms, continue incubating at room temperature. HINT for Step 7: To quickly confirm worm numbers, divide the plate into quarters. If you see 12 or more in the first quarter, the plate is ready.
59
1.800.EDVOTEK • www.edvotek.com • info@edvotek.com
Made with FlippingBook flipbook maker