Fig. 1: Knock-in of a ribozyme generates null-like mutants.

a Schematic of active T3H38 ribozyme (Rz) for inducing an off-switch of gene expression. Adapted with permission from Wurmthaler et al.¹¹. An inactive T3H38 (Rz*) was generated as a control. b Schematic diagram of CRISPR/Cas9-mediated homologous recombination to integrate GFP, GFP::Rz or GFP::Rz* at the C-terminal of endogenous loci. c Confocal fluorescent images to show the morphology of mitochondria (Pcol-19::mito-mKate), lysosomes (Pced-1::nuc-1::mCherry) and PVD (ser2prom::myr-mCherry) in related to the protein expression level of ARGN-1::GFP, SCAV-3::GFP and SAX-7::GFP, respectively, under different conditions. 3-day-old adults were imaged for the argn-1 and scav-3 groups; 2-day-old adults were imaged for the sax-7 group. Scale bars: 20 μm. Arrowheads: enlarged mitochondria/lysosome or dendrite branching defects. argn-1(zac427), scav-3(tm3659) and sax-7(nj48) are deletion mutants, resulting in strong loss-of-function/null alleles, and are used for comparisons. d Quantifications of the proportion of animals containing normal, intermediately defective and severely defective morphology of mitochondria (argn-1 group), lysosomes (scav-3 group) and PVD dendrites (sax-7 group), respectively. The number of animals quantified for each group was indicated above the columns. e Expression of ARGN-1::GFP, SCAV-3::GFP and SAX-7::GFP, as shown in (c), were quantified based on fluorescent imaging. Data are displayed as mean ± s.e.m. Each dot represents a single worm. Number of animals quantified: n = 35 for ARGN-1::GFP; n = 37 for ARGN-1::GFP::Rz*; n = 31 for ARGN-1::GFP::Rz; n = 41 for SCAV-3::GFP; n = 40 for SCAV-3::GFP::Rz*; n = 34 for SCAV-3::GFP::Rz; n = 41 for SAX-7::GFP; n = 32 for SAX-7::GFP::Rz*; n = 36 for SAX-7::GFP::Rz. ****p < 0.0001 (one-way ANOVA with the Tukey correction). f Relative mRNA abundance of target genes at different conditions in (c) measured by quantitative RT-PCR. Three biological replicates were quantified and shown. All values are displayed as mean ± s.e.m. Source data are provided as a Source Data file.