Collecting supernatant from C. elegans culture and lyophilizing supernatant
Muhammad Zaka Asif, Man Shah, Yosef Smadi
Abstract
This protocol describes day 5 of our workflow to grow worms in liquid culture to induce the production of natural products (in this case, 1-HP derivatives). In this protocol, we get an approximate count for the number of life-stage synchronized worms exposed to the toxin, and we collect the supernatant, which contains 1-HP and its derived metabolites. After collecting the supernatant, we freeze and lyophilize it to prepare for extraction.
Steps
Remove the flasks from the shaker and estimate their population by aliquoting onto glass slides as before.
Centrifuge at 525 RCF for 1 minute and collect the supernatant in multiple 2 mL microcentrifuge tubes.
Save the worm pellet at -80˚C if desired for future studies.
Centrifuge the microcentrifuge tubes at 20,800 RCF for 10 minutes. This will separate the bacteria from the rest of the supernatant (bacteria will settle to the bottom).
Combine all the supernatant remaining from the microcentrifuge tubes once the spin is completed in one 50 mL conical tube.
Freeze the supernatant with liquid nitrogen and lyophilize for 24-40 hrs until complete.
