WGS of C. elegans: ENU-treated morc-1 from pool 31-9
Starting with a morc-1(-);pkIs32(pie1P-gfp-h2b) strain, worms in the fourth larval stage were mutagenized in 0.6mM N-ethyl-N-nitrosourea (ENU) for 4 hours. After 3 washes in M9, 100 worms (P0 generation) were transferred to each of 32 plates. Worms we grown for 5 days at 20°C. Adult worms (F1 generation) were bleached to isolate embryos (F2 generation), which were plated, grown at 25°C, and then bleached to isolate embryos (F3 generation). Worms were maintained at 25°C and bleached at adulthood for every generation until F14. 10 larval worms were singled from each of the remaining fertile pools (29 of 32) and their progeny were counted. The most fertile worms of each pool were then propagated as clonalized lines. Genomic DNA was extracted from 20 clonalized lines representing 17 pools using the Gentra Puregene Tissue kit (Qiagen). Genomic DNA libraries were made using the Illumina Truseq Nano DNA LT Library Prep Kit according to manufacturer’s instructions.