pJW1138 (klp-12 targeting CRISPR/Cas9) and pJW1185 (nhr-25 targeting CRISPR/Cas9) were derived from pDD162 using a Q5 Mutagenesis kit (NEB, no. E0554S) as previously described (Dickinson et al. 2013 (link)). sgRNA(F+E) was synthesized as a gene fragment (IDT gBlock; sequence in Table S4) with a Y61A9LA.1 targeting sequence (Friedland et al. 2013 (link)) and introduced into a pDD162-derived vector by Gibson cloning (NEB, no. E5510S) to generate pJW1219. pJW1236 (klp-12 targeting CRISPR/Cas9), pJW1254 (nhr-23 PAM no. 1 targeting CRISPR/Cas9), pJW1268 (nhr-23 PAM no. 2 targeting CRISPR/Cas9) and pJW1285 (pha-1 targeting CRISPR/Cas9) were derived from pJW1219 [CRISPR/Cas9 with sgRNA(F+E)] through Q5 mutagenesis. The nhr-23, nhr-25, and pha-1 PAMs were manually chosen by searching for an NGG sequence in either strand close to the desired insertion site; these sgRNA target sites were then checked for specificity using the http://crispr.mit.edu website. All target sites scored >90 with no off-target sites in genes. The PU6::sgRNA template sequence was deleted from the pJW1219 vector using Q5 mutagenesis to generate pJW1259. All plasmids (standard vector propagation, and those generated by Gibson assembly, Q5 site-directed mutagenesis, or TOPO-blunt cloning) were transformed into PEG/DMSO DH5 alpha competent cells (protocol in File S1) made in house. pJW1219, pJW1259, pJW1285, pJW1310, and pJW1311 are available through AddGene.