GSM3861116: SOM RNA-seq rep1; Caenorhabditis elegans; RNA-Seq
Growth:  glp-1(q224) animals were cultured to starvation on NGM plates at 15Â°C. L1 worms were floated and plated on one 15-cm NGM plate. Worms were cultured at 15Â°C for four days until gravid. Adult worms were bleached and embryos were incubated with shaking at 15Â°C for 36-42 hours. 15K L1s were plated to one peptone enriched plate and cultured at 25Â°C for 46-48 hours until adult stage. Extraction: Worms were harvested by washing with M9 3x and were incubated with shaking for 30â??45 minutes in M9 buffer. Worms were centifuged at 3100 rpm for 2 minutes. Supernatant was removed and 500 ÂµL TRIzol was added to the worm pellet.  Total RNA isolation was performed with standard TRIzol RNA extraction. Total RNA was then treated with DNA-free rDNase I and cleaned up using RNeasy Mini Kit. rRNA was depleted by Ribo-Zero rRNA Removal Kit. Library construction:  The Yale Center for Genome Analysis (YCGA) prepared libraries for each sample using the Kapa Biosystems reagents.