CRISPR co-editing protocol for odr-3 mutant

The odr-3(rms31) mutant was generated by CRISPR using a dual crRNA dpy-10 co-CRISPR strategy and a custom protocol based on previous methods^{52 (link),53 (link)} and optimization for our laboratory. In brief, 1 µl of 320 µM solution of each CRISPR RNA (crRNA) and 0.5 µl of dpy-10 crRNA (50 µM) was annealed to 0.4 µl of 100 µM trans-acting CRISPR RNA (tracrRNA) (Integrated DNA Technologies) by heating to 95 °C in a PCR machine and cooling to 4 °C at 0.1 °C s⁻¹. Then, 0.5 µl of Cas9 protein (Invitrogen) was added, and the mixture was incubated for 10 min at 37 °C. Next, 0.5 µl of 100 µM stock of each repair template (target and dpy-10) was added, and the solution was made up to 20 µl with DPEC water. This mix was centrifuged for 30 min at 20,000g at 4 °C before injection. Oligonucleotides used in this study are provided in Supplementary Tables 3 and 4.