4.7. Gene Expression Assay

Total RNA was extracted from C. elegans with the Trizol reagent (Invitrogen, Waltham, MA, USA). Reverse transcriptase reactions required 1 μg RNA to synthesize complementary cDNA utilizing a Reverse Transcription kit (Invitrogen, USA) according to the manufacturer’s protocol. Incubation conditions included 10 min at 25 °C, 120 min at 37 °C and 5 min at 85 °C. The resulting cDNAs were used to identify the level of serotonin-related gene expression (mod-1, mod-5, cat-4, tph-1) and stress-related gene expression (sod-1, sod-3, ctl-2, ctl-3, mtl-1, cyp) by quantitative PCR using SybrGreen PCR Master Mix Kit (Applied Biosystems, Waltham, MA, USA) and specific primers as previously described [11].