Quantification of C. elegans Yolk Proteins

Coomassie staining of worm protein extracts using SDS-PAGE was performed as described in [3 (link),27 (link)]. Synchronous worm populations were grown until day 3 of adulthood, after which 50 animals per sample were picked into 15 μL of M9 buffer. A total of 15 μL of Laemmli sample buffer containing β-mercaptoethanol was added. Samples were incubated for 15 minutes at 70°C, centrifuged at max speed for 5 minutes, and incubated at 95°C for 5 minutes. A total of 15 μL of each sample was loaded on a 4%–12% bis-tris Criterion XT precast polyacrylamide gel (Bio-Rad, Hercules, CA) using XT MOPS as a running buffer. Gels were stained with Coomassie Brilliant Blue and destained with a 40% methanol, 10% acetic acid solution. Gel images were taken with a Bio-Rad Gel Doc (Bio-Rad, Hercules, CA). Identification of YP170, YP115, and YP88 bands is based on [7 (link),27 (link)]. Quantification of yolk protein abundance was done by normalizing to total protein content in a lane using ImageLab 6.0. At least three populations were assayed for each condition. Data were analyzed using a one-way ANOVA with a Tukey post hoc test.