Feeding RNAi protocol

Day 1: Day 2: Day 3: Day 6 or 7:
 * Start 20ml LB + carbenicillin cultures of your bacterial strain containing the RNAi vector.
 * Don’t forget your empty vector control (L4440)!
 * Grow overnight at 37º.
 * Chunk worms and put at 20°
 * In the morning, spin down the overnight cultures (5 min at 4,000 rpm in the table top centrifuge works), and resuspend them in 1ml total volume. This stock is enough for 10 plates and can be kept for up to a week at 4º.
 * Put 100µl/plate of the resuspended bacteria onto NGM + IPTG + carbenicillin
 * After the plates have dried, put plates at 37º to induce RNA expression overnight
 * The next morning, put ~100µl of M9 onto an unspread NGM plate. You can also use the cover of one of your RNAi plates. Pick wildtype L4s into the M9 to rinse. You can pipette up and down if you want.
 * Pick L4s from the M9 and put them on one of the plates seeded the day before with RNAi bacteria. You can also pipette them into the plate and let it dry, assuming that there is not very much OP50 in the M9. Put them at 20º and let them crawl around eating the bacteria for ~2 hrs. This should get rid of any residual OP50 in their gut.
 * After 2 hrs, transfer the worms to fresh plates (seeded the day before with RNAi bacteria). Put 3-4 worms/plate and keep at 20º to let the parents self. Put the remaining plates at 4°.
 * If your experiment requires staged worms (i.e. immunofluorescence or scoring apoptosis), pick L4s to the RNAi plate stored at 4° and put at 20º to perform the experiment the next day.