Table 3:

Troubleshooting.

Problem	Possible Causes	Solutions
No phenotype	• RNAi didn’t work • RNAi food contaminated	• Prepare fresh media and IPTG • Use fresh bacterial culture for induction • Test the assay with positive controls • Make fresh antibiotics and store them at an appropriate temperature • Use carbenicillin instead of ampicillin
Edge Effect	• The wells in edge dry faster than the wells in the center	• Incubate the plate in a well-humidified chamber or box • If using 96 well plates, shift to 96 half-area well plates for the screen
The image has lots of debris and background	• Worm debris, salts in the buffer, and bacteria can increase background	• Washing with M9 or S- basal will reduce the debris and will give a clean image for analysis
Images with lots of wiggling worms	• Worms tend to swim more actively under the microscope when illuminated with blue light	• Add levamisole or keep the worms on ice for few minutes before imaging.