Whole genome sequencing of C. elegans parental control for comparison
ns942 and parental OS12214 animals were grown on E. coli strain OP50 until bacteria were depleted, harvested in M9, and resuspended in 0.5mL TEN buffer (20 mm Tris pH 7.5, 0.5 M EDTA, 100 mM NaCl), pH 7.5. SDS (0.5%), proteinase K (0.1 mg/mL), and -mercaptoethanol (0.2%) were added. The lysis reaction was incubated overnight in a shaking thermocycler at 56C. Phenol/chloroform was added and phase-separated by spinning in a phase-lock tube. The aqueous phase was transferred to 200 proof EtOH. The resulting DNA clot was washed in 70% EtOH, dried, and resuspended in TEN. After rehydration, 0.3 L of 100 mg/mL RNAse A was added and incubated at 37C for 2 h. Phenol/chloroform extraction was performed again and DNA was rehydrated with EB buffer. The sample was then examined using a nanodrop spectrophotometer and run on a 1% agarose gel to confirm that there was no RNA contamination or DNA degradation, respectively. NextSeq Mid Output 2 x 150 sequencing was performed using Standard Illumina Sequencing primers for gDNA-seq application.